Pharmacologically active agents containing esterified phosphonates and methods for use thereof

ABSTRACT

Compounds and compositions are provided for treatment, prevention, or amelioration of a variety of medical disorders associated with viral infections, cell proliferation and bone metabolism. The compounds provided herein are alkyl esters of phosphonates.

RELATED APPLICATION DATA

This application claims priority under 35 U.S.C. §119(e) to U.S.provisional application Ser. No. 60/542,522, filed Feb. 5, 2004,entitled “PHARMACOLOGICALLY ACTIVE AGENTS CONTAINING ESTERIFIEDPHOSPHONATES AND METHODS FOR USE THEREOF” to Karl Hostetler and BradWan, which is incorporated herein by reference in its entirety.

GRANT INFORMATION

This invention was made with government support under Grant No.EY11832-06 awarded by the National Health Institute/National EyeInstitute, and under Grant No. DAMD17-01-2-0071 awarded by theDepartment of Defense. The United States government has certain rightsin this invention.

FIELD

Provided herein are alkyl esters of phosphonate compounds. In oneembodiment, the compounds are esterified derivatives of biologicallyactive phosphonates. In another embodiment, provided herein are methodsof treatment, prevention, or amelioration of a variety of medicaldisorders associated with viral infections, cell proliferation and bonemetabolism using the compounds and compositions provided herein.

BACKGROUND

Phosphonate compounds have long been known to have antiviral,antiproliferative and other variety of therapeutic benefits. Among theseare the antiviral nucleotide phosphonates, such as, for example,cidofovir, cyclic cidofovir, adefovir, tenofovir, and the like, as wellas the 5′-phosphonates and methylene phosphonates of azidothymidine(AZT), ganciclovir, acyclovir, and the like. In these compounds, the5′-hydroxyl of the sugar moiety, or its equivalent in acyclicnucleosides (ganciclovir, penciclovir, acyclovir) which do not contain acomplete sugar moiety, is replaced with a phosphorus-carbon bond. In thecase of the methylene phosphonates, a methylene group replaces the5′-hydroxyl or its equivalent, and its carbon atom is, in turn,covalently linked to the phosphonate.

Such compounds may be active as antiviral or antiproliferativenucleotides. Upon cellular metabolism, two additional phosphorylationsoccur to form the nucleotide phosphonate diphosphate which representsthe equivalent of nucleoside triphosphates. Antiviral nucleotidephosphonate diphosphates are selective inhibitors of viral RNA or DNApolymerases or reverse transcriptases. That is to say, their inhibitoryaction on viral polymerases is much greater than their degree ofinhibition of mammalian cell DNA polymerases α, β and γ or mammalian RNApolymerases. Conversely, the antiproliferative nucleotide phosphonatediphosphates inhibit cancer cell DNA and RNA polymerases and may showmuch lower selectivity versus normal cellular DNA and RNA polymerases.

Another class of therapeutically beneficial phosphonate compounds arethe bisphosphonates, i.e., pyrophosphate analogs wherein the centraloxygen atom of the pyrophosphate bond is replaced by carbon. Varioussubstituent groups may be attached to this central carbon atom toproduce derivatives of bisphosphonate compounds having various degreesof pharmacological potency.

Bisphosphonates and their substituted derivatives have the intrinsicproperty of inhibiting bone resorption in vivo. Bisphosphonates also actby inhibiting apoptosis (programmed cell death) in bone-forming cells.Indications for their use therefore include the treatment and preventionof osteoporosis, treatment of Paget's disease, metastatic bone cancers,hyperparathyroidism, rheumatoid arthritis, algodistrophy,sterno-costo-clavicular hyperostosis, Gaucher's disease, Engleman'sdisease, and certain non-skeletal disorders.

Although bisphosphonates have therapeutically beneficial properties,they suffer from pharmacological disadvantages as orally administeredagents. One drawback is low oral availability: as little as 0.7% to 5%of an orally administered dose is absorbed from the gastrointestinaltract. Oral absorption is further reduced when taken with food. Further,it is known that some currently available bisphosphonates, e.g.,FOSAMAX® (Merck; alendronate sodium), SKELID® (Sanofi, tiludronate) andACTONEL® (Procter and Gamble, risedronate) have local toxicity, causingesophageal irritation and ulceration. Other bisphosphonates, likeamino-olpadronate, lack anti-resorptive effects (Van Beek, E. et al., J.Bone Miner Res 11(10): 1492-1497 (1996) but inhibit osteocyte apoptosisand are able to stimulate net bone formation (Plotkin, L. et al., J ClinInvest 104(10):1363-1374 (1999) and U.S. Pat. No. 5,885,973). It wouldtherefore, be useful to develop chemically modified bisphosphonatederivatives that maintain or enhance the pharmacological activity of theparent compounds while eliminating or reducing their undesirable sideeffects.

The threat of an intentional or an unintentional spread of poxvirusinfections to a vulnerable population has led to increased efforts tofind safe, rapidly deployable treatments against such infections.Although vaccination is now being offered to some healthcare workers andother first responders, there are valid concerns about potential vaccinerisks. Previously reported smallpox vaccine-associated adversereactions, along with the unknown prevalence of risk factors amongtoday's population has prompted the preparation of guidances forclinicians in evaluating and treating patients with smallpox vaccinationcomplications. Following this guidance, the vaccine is not recommendedfor those with eczema and other exfoliative skin disorders, those withhereditary or acquired immunodeficiencies, or for pregnant women orwomen who desire to become pregnant 28 days after vaccination. Morerecently the Centers for Disease Control issued a health advisoryrecommending as a precautionary measure that persons with known cardiacdisease not be vaccinated at this time. Therefore, the use of antiviraltherapy in the event of a poxvirus outbreak or in the treatment ofvaccination complications against smallpox virus points to the continuedneed to examine available antiviral therapies, as well as thedevelopment of new and more efficient treatment regimens.

There is, therefore, a continuing need for less toxic, more effectivepharmaceutical agents to treat a variety of disorders associated withviral infection, cell proliferation and bone metabolism.

SUMMARY

Provided herein are phosphonate and bisphosphonate alkyl esters andcompositions thereof for the treatment of various diseases. In oneembodiment, compounds and compositions provided herein have antiviralactivity. In another embodiment, provided herein are compounds andcompositions that are useful in the treatment, prevention, oramelioration of one or more symptoms associated with cell proliferation.In yet another embodiment, the compounds and compositions are fortreating diseases associated with bone metabolism in a subject.

Provided herein are chemically modified phosphonate derivatives ofpharmacologically active agents, e.g., antiviral and anti-neoplasticpharmaceutical agents that contain phosphonates. These modifiedderivatives increase the potency of the parent compound while minimizingdeleterious side effects when administered to a subject in need thereof.

In certain embodiments, the compounds are lipophilic esters ofphosphonates. In certain embodiments, the lipophilic esters exhibitenhanced activity against cells infected with poxviruses and herpesviruses compared to non-esterified phosphonates.

In one embodiment, the compounds for use in the compositions and methodsprovided herein have formula I:P^(x)—O—R

wherein P^(x) is a pharmacologically active phosphonate, O is an oxygenatom, and R is a substituted or unsubstituted C8-C24 alkyl, substitutedor unsubstituted C8-C24 alkenyl having from 1 to 6 double bonds orsubstituted or unsubstituted C8-C24 alkynyl having from 1 to 6 triplebonds, wherein substituents when present are selected from one or morehalogen, alkyl, —OR^(w), —SR^(w), cycloalkyl or epoxide, where R^(w) ishydrogen or alkyl and where the alkyl, alkenyl, alkynyl groups may befurther substituted or unsubstituted.

Also provided are pharmaceutically-acceptable derivatives, includingsalts, esters, enol ethers, enol esters, solvates, hydrates and prodrugsof the compounds described herein. Further provided are pharmaceuticalcompositions containing the compounds provided herein and apharmaceutically acceptable carrier. In one embodiment, thepharmaceutical compositions are formulated for single dosageadministration.

Methods of treating, using the compounds and compositions providedherein are provided. Methods of treating, preventing, or amelioratingone or more symptoms of diseases associated with viral infections, cellproliferation and bone metabolism using the compounds and compositionsprovided herein are provided. In practicing the methods, effectiveamounts of the compounds or compositions containing therapeuticallyeffective concentrations of the compounds are administered.

Articles of manufacture are provided containing packaging material, acompound or composition provided herein which is useful for treating,preventing, or ameliorating one or more symptoms of diseases ordisorders associated with viral infections, cell proliferation or bonemetabolism using the compounds and compositions provided herein, and alabel that indicates that the compound or composition is useful fortreating, preventing, or ameliorating one or more symptoms of diseasesor disorders associated with viral infections, cell proliferation orbone metabolism.

DETAILED DESCRIPTION A. Definitions

Unless defined otherwise, all technical and scientific terms used hereinhave the same meaning as is commonly understood by one of ordinary skillin the art to which this invention belongs. All patents, applications,published applications and other publications are incorporated byreference in their entirety. In the event that there are a plurality ofdefinitions for a term herein, those in this section prevail unlessstated otherwise.

Phosphonate compounds for use herein are biologically active derivativesof phosphonic acid that can be converted into alkyl esters as describedherein. In certain embodiments, the phosphonate compounds for use hereinare phosphonate-containing nucleotides or nucleosides which can bederivatized to their corresponding phosphonates. In other embodiments,the phosphonate compounds for use herein are bisphosphonates.

Bisphosphonates are synthetic phosphonic acid derivatives characterizedby two carbon-phosphorus bonds. There are no known enzymes that cancleave such bonds with the consequence that bisphosphonates areabsorbed, stored and excreted from the body unaltered. Theirphysiochemical effect is similar to that of pyrophosphate in that theybind strongly to the surface of solid-phase calcium phosphate and bydoing so inhibit the formation, delay the aggregation and slow down thedissolution of calcium phosphate crystals.

As used herein, pharmaceutically acceptable derivatives of a compoundinclude salts, esters, enol ethers, enol esters, acetals, ketals,orthoesters, hemiacetals, hemiketals, acids, bases, solvates, hydratesor prodrugs thereof. Such derivatives may be readily prepared by thoseof skill in this art using known methods for such derivatization. Thecompounds produced may be administered to animals or humans withoutsubstantial toxic effects and either are pharmaceutically active or areprodrugs. Pharmaceutically acceptable salts include, but are not limitedto, amine salts, such as but not limited toN,N′-dibenzylethylenediamine, chloroprocaine, choline, ammonia,diethanolamine and other hydroxyalkylamines, ethylenediamine,N-methylglucamine, procaine, N-benzylphenethylamine,1-para-chlorobenzyl-2-pyrrolidin-1′-ylmethyl-benzimidazole, diethylamineand other alkylamines, piperazine and tris(hydroxymethyl)aminomethane;alkali metal salts, such as but not limited to lithium, potassium andsodium; alkali earth metal salts, such as but not limited to barium,calcium and magnesium; transition metal salts, such as but not limitedto zinc; and other metal salts, such as but not limited to sodiumhydrogen phosphate and disodium phosphate; and also including, but notlimited to, nitrates, borates, methanesulfonates, benzenesulfonates,toluenesulfonates, salts of mineral acids, such as but not limited tohydrochlorides, hydrobromides, hydroiodides and sulfates; and salts oforganic acids, such as but not limited to acetates, trifluoroacetates,maleates, oxalates, lactates, malates, tartrates, citrates, benzoates,salicylates, ascorbates, succinates, butyrates, valerates and fumarates.Pharmaceutically acceptable esters include, but are not limited to,alkyl, alkenyl, alkynyl, and cycloalkyl esters of acidic groups,including, but not limited to, carboxylic acids, phosphoric acids,phosphinic acids, sulfonic acids, sulfinic acids and boronic acids.Pharmaceutically acceptable enol ethers include, but are not limited to,derivatives of formula C═C(OR) where R is hydrogen, alkyl, alkenyl,alkynyl, and cycloalkyl. Pharmaceutically acceptable enol estersinclude, but are not limited to, derivatives of formula C═C(OC(O)R)where R is hydrogen, alkyl, alkenyl, alkynyl, or cycloalkyl.Pharmaceutically acceptable solvates and hydrates are complexes of acompound with one or more solvent or water molecules, or 1 to about 100,or 1 to about 10, or one to about 2, 3 or 4, solvent or water molecules.

As used herein, treatment means any manner in which one or more of thesymptoms of a disease or disorder are ameliorated or otherwisebeneficially altered. Treatment also encompasses any pharmaceutical useof the compositions herein, such as use for treating diseases ordisorders in which α-synuclein fibril formation is implicated.

As used herein, amelioration of the symptoms of a particular disorder byadministration of a particular compound or pharmaceutical compositionrefers to any lessening, whether permanent or temporary, lasting ortransient that can be attributed to or associated with administration ofthe composition.

As used herein, EC₅₀ refers to a dosage, concentration or amount of aparticular test compound that elicits a dose-dependent response at 50%of maximal expression of a particular response that is induced, provokedor potentiated by the particular test compound.

As used herein, a prodrug is a compound that, upon in vivoadministration, is metabolized by one or more steps or processes orotherwise converted to the biologically, pharmaceutically ortherapeutically active form of the compound. To produce a prodrug, thepharmaceutically active compound is modified such that the activecompound will be regenerated by metabolic processes. The prodrug may bedesigned to alter the metabolic stability or the transportcharacteristics of a drug, to mask side effects or toxicity, to improvethe flavor of a drug or to alter other characteristics or properties ofa drug. By virtue of knowledge of pharmacodynamic processes and drugmetabolism in vivo, those of skill in this art, once a pharmaceuticallyactive compound is known, can design prodrugs of the compound (see,e.g., Nogrady (1985) Medicinal Chemistry A Biochemical Approach, OxfordUniversity Press, New York, pages 388-392). Other prodrugs for useherein are described elsewhere herein.

It is to be understood that the compounds provided herein may containchiral centers. Such chiral centers may be of either the (R) or (S)configuration, or may be a mixture thereof. Thus, the compounds providedherein may be enantiomerically pure, or be stereoisomeric ordiastereomeric mixtures. It is understood that the present inventionencompasses any racemic, optically active, polymorphic, orstereoisomeric form, or mixtures thereof, of a compound of theinvention, which possesses the useful properties described herein, itbeing well known in the art how to prepare optically active forms andhow to determine antiproliferative activity using the standard testsdescribed herein, or using other similar tests which are will known inthe art. Examples of methods that can be used to obtain optical isomersof the compounds of the present invention include the following:

i) physical separation of crystals—a technique whereby macroscopiccrystals of the individual enantiomers are manually separated. Thistechnique can be used if crystals of the separate enantiomers exist,i.e., the material is a conglomerate, and the crystals are visuallydistinct;

ii) simultaneous crystallization—a technique whereby the individualenantiomers are separately crystallized from a solution of the racemate,possible only if the latter is a conglomerate in the solid state;

iii) enzymatic resolutions—a technique whereby partial or

complete separation of a racemate by virtue of differing rates ofreaction for the enantiomers with an enzyme

iv) enzymatic asymmetric synthesis—a synthetic technique

whereby at least one step of the synthesis uses an enzymatic reaction toobtain an enatiomerically pure or enriched synthetic precursor of thedesired enantiomer;

v) chemical asymmetric synthesis—a synthetic technique whereby thedesired enantiomer is synthesized from an achiral precursor underconditions that produce assymetry (i.e., chirality) in the product,which may be achieved using chiral catalysts or chiral auxiliaries;

vi) diastereomer separations—a technique whereby a racemic

compound is reacted with an enantiomerically pure reagent (the chiralauxiliary) that converts the individual enantiomers to diastereomers.The resulting diastereomers are then separated by chromatography orcrystallization by virtue of their now more distinct structuraldifferences and the chiral auxiliary later removed to obtain the desiredenantiomer;

vii) first- and second-order asymmetric transformations—a

technique whereby diastereomers from the racemate equilibrate to yield apreponderance in solution of the diastereomer from the desiredenantiomer or where preferential crystallization of the diastereomerfrom the desired enantiomer perturbs the equilibrium such thateventually in principle all the material is converted to the crystallinediastereomer from the desired enantiomer. The desired enantiomer is thenreleased from the diastereomer;

viii) kinetic resolutions—this technique refers to the

achievement of partial or complete resolution of a racemate (or of afurther resolution of a partially resolved compound) by virtue ofunequal reaction rates of the enantiomers with a chiral, non-racemicreagent or catalyst under kinetic conditions;

ix) enantiospecific synthesis from non-racemic precursors—a synthetictechnique whereby the desired enantiomer is obtained from non-chiralstarting materials and where the stereochemical integrity is not or isonly minimally compromised over the course of the synthesis;

x) chiral liquid chromatography—a technique whereby the enantiomers of aracemate are separated in a liquid mobile phase by virtue of theirdiffering interactions with a stationary phase. The stationary phase canbe made of chiral material or the mobile phase can contain an additionalchiral material to provoke the differing interactions;

xi) chiral gas chromatography—a technique whereby the racemate isvolatilized and enantiomers are separated by virtue of their differinginteractions in the gaseous mobile phase with a column containing afixed non-racemic chiral adsorbent phase;

xii) extraction with chiral solvents—a technique whereby the enantiomersare separated by virtue of preferential dissolution of one enantiomerinto a particular chiral solvent;

xiii) transport across chiral membranes—a technique whereby a racemateis placed in contact with a thin membrane barrier. The barrier typicallyseparates two miscible fluids, one containing the racemate, and adriving force such as concentration or pressure differential causespreferential transport across the membrane barrier. Separation occurs asa result of the non-racemic chiral nature of the membrane which allowsonly one enantiomer of the racemate to pass through.

In the case of amino acid residues, such residues may be of either theL- or D-form. The configuration for naturally occurring amino acidresidues is generally L. When not specified the residue is the L form.As used herein, the term “amino acid” refers to α-amino acids which areracemic, or of either the D- or L-configuration. The designation “d”preceding an amino acid designation (e.g., dAla, dSer, dVal, etc.)refers to the D-isomer of the amino acid. The designation “dl” precedingan amino acid designation (e.g., dlPip) refers to a mixture of the L-and D-isomers of the amino acid. It is to be understood that the chiralcenters of the compounds provided herein may undergo epimerization invivo. As such, one of skill in the art will recognize thatadministration of a compound in its (R) form is equivalent, forcompounds that undergo epimerization in vivo, to administration of thecompound in its (S) form.

As used herein, substantially pure means sufficiently homogeneous toappear free of readily detectable impurities as determined by standardmethods of analysis, such as thin layer chromatography (TLC), gelelectrophoresis, high performance liquid chromatography (HPLC) and massspectrometry (MS), used by those of skill in the art to assess suchpurity, or sufficiently pure such that further purification would notdetectably alter the physical and chemical properties, such as enzymaticand biological activities, of the substance. Methods for purification ofthe compounds to produce substantially chemically pure compounds areknown to those of skill in the art. A substantially chemically purecompound may, however, be a mixture of stereoisomers. In such instances,further purification might increase the specific activity of thecompound.

As used herein, the term “alkyl” refers to a monovalent straight orbranched chain or cyclic radical. In certain embodiments, the alkylgroup contains from one to twenty-four carbon atoms, including methyl,ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, n-hexyl,octadecyl, nonadecyl, eicosyl, 18-methyl-nonadecyl, 19-methyl-eicosyl,and the like. As used herein lower alkyl refers to alkyl groups of 1 to6 carbon atoms.

As used herein, “substituted alkyl” refers to alkyl groups furtherbearing one or more substituents, including, but not limited tosubstituents selected from lower alkyl, hydroxy, alkoxy (of a loweralkyl group), mercapto (of a lower alkyl group), cycloalkyl, substitutedcycloalkyl, heterocyclic, substituted heterocyclic, aryl, substitutedaryl, heteroaryl, substituted heteroaryl, aryloxy, substituted aryloxy,halogen, trifluoromethyl, cyano, azido, nitro, nitrone, amino, amido,—C(O)H, acyl, oxyacyl, carboxyl, carbamate, sulfonyl, sulfonamide, andsulfuryl, which may be protected or unprotected as necessary, as taughtin Greene, et al., Protective Groups in Organic Synthesis, John Wileyand Sons, Second Ed. 1991, hereby incorporated by reference.

As used herein, “alkenyl” refers to straight or branched chainhydrocarbon group having one or more carbon-carbon double bonds. Incertain embodiments, the alkenyl group contains from 2 up to 24 carbonatoms, and “substituted alkenyl” refers to alkenyl groups furtherbearing one or more substituents as set forth above.

As used herein, “alkynyl” refers to straight or branched chainhydrocarbon group having one or more carbon-carbon triple bonds. Incertain embodiments, the alkynyl group contains from 2 up to 24 carbonatoms, and “substituted alkynyl” refers to alkynyl groups furtherbearing one or more substituents as set forth above.

As used herein, “aryl” refers to aromatic groups having in the range of6 up to 14 carbon atoms and “substituted aryl” refers to aryl groupsfurther bearing one or more substituents as set forth above.

As used herein, “heteroaryl” refers to aromatic groups containing one ormore heteroatoms (e.g., N, O, S, or the like) as part of the ringstructure, and having in the range of 3 up to 14 carbon atoms and“substituted heteroaryl” refers to heteroaryl groups further bearing oneor more substituents as set forth above.

The term “nucleoside” as used herein, refers to a molecule composed of aheterocyclic base and a carbohydrate. Typically, a nucleoside iscomposed of a heterocyclic nitrogenous base in N-glycosidic linkage witha sugar. Nucleosides are recognized in the art to include natural bases(standard), and non-natural bases well known in the art. Thecarbohydrates include the true sugars found in natural nucleosides or aspecies replacing the ribofuranosyl moiety or acyclic sugars. Theheterocyclic nitrogenous bases are generally located at the 1′ positionof a nucleoside sugar moiety. Nucleosides generally contain a base andsugar group. The nucleosides can be unmodified or modified at the sugar,and/or base moiety, (also referred to interchangeably as nucleosideanalogs, modified nucleosides, non-natural nucleosides, non-standardnucleosides; see for example, Eckstein et al., International PCTPublication No. WO 92/07065 and Usman et al., International PCTPublication No. WO 93/15187). In natural nucleosides the heterocyclicbase is typically thymine, uracil, cytosine, adenine or guanine. Thecarbohydrate shall be understood to mean the true sugar found in naturalnucleosides or a species replacing the ribofuranosyl moiety or acyclicsugars. In certain embodiments, acyclic sugars contain 3-6 carbon atomsand include, for example, the acyclic sugar moieties present inacyclovir (—CH2-O—CH2CH2-OH), ganciclovir (—CH2-O—CH(CH2OH)—CH2-OH), andthe like. Natural nucleosides have the β-D-configuration. The term“nucleoside” shall be understood to encompass unnatural configurationsand species replacing the true sugar that lack an anomeric carbon. Innatural nucleosides the heterocyclic base is attached to thecarbohydrate through a carbon-nitrogen bond. The term “nucleoside” shallbe understood to encompass species wherein the heterocyclic base andcarbohydrate are attached through a carbon-carbon bond (C-nucleosides).

As used herein “subject” is an animal, typically a mammal, includinghuman, such as a patient.

The phrase “effective amount” as used herein means an amount requiredfor prevention, treatment, or amelioration of one or more of thesymptoms of diseases or disorders associated including those associatedwith viral infection, cell proliferation and/or bone metabolism.

Where the number of any given substituent is not specified (e.g.,haloalkyl), there may be one or more substituents present. For example,“haloalkyl” may include one or more of the same or different halogens.

As used herein, the term “parenteral” includes subcutaneous,intravenous, intra-arterial, intramuscular or intravitreal injection, orinfusion techniques.

The term “topically” encompasses administration rectally and byinhalation spray, as well as the more common routes of the skin andmucous membranes of the mouth and nose and in toothpaste.

As used herein, the abbreviations for any protective groups, amino acidsand other compounds, are, unless indicated otherwise, in accord withtheir common usage, recognized abbreviations, or the IUPAC-IUBCommission on Biochemical Nomenclature (see, (1972) Biochem.11:942-944).

B. Compounds

In certain embodiments, provided herein are alkyl ester derivatives ofknown phosphonate compounds according to the methods provided herein. Incertain embodiments, the alkyl ester derivatives of phosphonatesprovided herein have improved properties, such as improved pharmacologicactivity, or increased oral absorption. In certain embodiments, cellularenzymes, but not plasma or digestive tract enzymes, convert thecompounds provided herein to a free phosphonates. In certainembodiments, the phosphonate compounds described herein reduce oreliminate the tendency of co-administered food to reduce or abolishphosphonate absorption, resulting in higher plasma levels ofphosphonates and better compliance by patients.

In one embodiment, the alkyl ester compounds for use in the compositionsand methods provided herein have formula I:P^(x)—O—R

wherein P^(x) is a pharmacologically active phosphonate, O is an oxygenatom, and R is a substituted or unsubstituted C₈-C₂₄ alkyl, substitutedor unsubstituted C₈-C₂₄ alkenyl having from 1 to 6 double bonds orsubstituted or unsubstituted C₈-C₂₄ alkynyl having from 1 to 6 triplebonds wherein substituents when present are selected from one or morehalogen, alkyl, alkenyl, alkynyl, —OR^(w), —SR^(w), cycloalkyl orepoxide, where each R^(w) is independently hydrogen or alkyl and wherethe alkyl, alkenyl, alkynyl groups may be substituted or unsubstituted.

In one embodiment, the compounds for use in the compositions and methodsprovided herein have formula I:P^(x)—O—R

wherein P^(x) is a pharmacologically active phosphonate, O is an oxygenatom, and R is a substituted or unsubstituted C₈-C₂₄ alkyl orsubstituted or unsubstituted C₈-C₂₄ alkenyl having from 1 to 6 doublebonds, wherein substituents when present are selected from one or morehalogen, alkyl, —OH, —SH, cycloalkyl, or epoxide.

In certain embodiments, the R group in the alkyl, alkenyl and alkynylgroups in the compounds of formula I contain 8, 10, 12, 13, 14, 15, 16,17, 18, 19, 20, 21, 22, 23 or 24 carbon atoms and can be straight orbranched chain moieties. In certain embodiments, the R group is aC₁₆-C₂₃ straight or branched chain alkyl or C₁₆-C₂₃ straight or branchedchain alkenyl. In other embodiments, R is a C₁₇-C₁₉ straight or branchedchain alkyl or C₁₇-C₁₉ straight or branched chain alkenyl. In otherembodiments, R is C₁₇-alkyl, C₁₈-alkyl or C₁₉ alkyl. In otherembodiments, R is C₁₇-alkenyl, C₁₈-alkenyl or C₁₉ alkenyl. In otherembodiments, R is C₁₇-C₂₂ alkyl. In other embodiments, R is C₁₇ alkyl,C₁₈ alkyl, C₁₉ alkyl, C₂₀ alkyl, C₂₁ alkyl, or C₂₂ alkyl.

In certain embodiments, R is substituted with one or more groupsselected from lower alkyl and halo. In certain embodiments, R issubstituted with one or more methyl groups. In certain embodiments, R issubstituted with one or more fluoro groups. In certain embodiments, R isC₁₆-C₂₃ alkyl and is substituted with one or more methyl or fluorogroups. In certain embodiments, the methyl group or the fluoro groupsubstituent is present on the penultimate carbon of the alkyl, alkenyl,or alkynyl chain. In certain embodiments, the R is 7-methyl-octyl,8-methyl-nonyl, 9-methyl-decyl, 10-methyl-undecyl, 11-methyl-dodecyl,12-methyl-tridecyl, 13-methyl-tetradecyl, 14-methyl-pentadecyl,15-methyl-hexadecyl, 16-methyl-heptadecyl, 17-methyl-octadecyl,18-methyl-nonadecyl, 19-methyl-eicosyl, 20-methyl-heneicosyl,21-methyl-docosyl, 22-methyl-tricosyl, 7-fluoro-octyl, 8-fluoro-nonyl,9-fluoro-decyl, 10-fluoro-undecyl, 11-fluoro-dodecyl,12-fluoro-tridecyl, 13-fluoro-tetradecyl, 14-fluoro-pentadecyl,15-fluoro-hexadecyl, 16-fluoro-heptadecyl, 17-fluoro-octadecyl,18-fluoro-nonadecyl, 19-fluoro-eicosyl, 20-fluoro-heneicosyl,21-fluoro-docosyl or 22-fluoro-tricosyl.

Various pharmacologically active phosphonates can be used in thecompounds of formula I provided herein. In certain embodiments, P^(x) isrepresented by formula II

wherein R₁ is an antiviral nucleoside or antiproliferative nucleoside,or a bisphosphonate, and R′ is selected from an —H, a substituted orunsubstituted straight chain, branched or cyclic C8-C24 alkyl,substituted or unsubstituted C8-C24 alkenyl or C8-C24 alkynyl andwherein substituents when present are selected from one or more halogen,alkyl, alkenyl, alkynyl —OR₃, —SR₃, cycloalkyl, or epoxide and R₃ is a—H or lower alkyl; is —H, or a physiologically acceptable monovalentcation.

Various pharmacologically active phosphonates can be used in thecompounds of formula I provided herein. In certain embodiments, P^(x) isrepresented by formula II

wherein R₁ is an antiviral nucleoside or antiproliferative nucleoside,or a bisphosphonate, and M is —H, a physiologically acceptablemonovalent cation, a substituted or unsubstituted straight chain,branched or cyclic C8-C24 alkyl, substituted or unsubstituted C8-C24alkenyl or C8-C24 alkynyl and wherein substituents when present areselected from one or more halogen, alkyl, alkenyl, alkynyl, OR^(w),—SR^(w), cycloalkyl or epoxide, where each R^(w) is independentlyhydrogen or C1-6 alkyl. In certain embodiments, M is —H, or aphysiologically acceptable monovalent cation.

In certain embodiments, P^(x) is represented by formula II

wherein R₁ is an antiviral nucleoside or antiproliferative nucleoside,or a bisphosphonate, and M is —H, Na⁺, K⁺, or a physiologicallyacceptable monovalent cation.

In certain embodiments, the compounds provided herein have formula:

or a pharmaceutically acceptable derivative thereof,wherein R³, R⁴ and R⁵ are each H, hydroxy, halo, azido, C1-6 alkyl, C2-6alkenyl or C2-6 alkynyl; B is a purine or pyrimidine base or analogthereof and P¹ is

where M and R are as defined elsewhere herein. In certain embodiments,R⁴ and R⁵ are selected from hydrogen, halo and hydroxyalkyl. In certainembodiments, R⁴ and R⁵ are selected from halo and hydroxyalkyl. Incertain embodiments, R⁴ and R⁵ are selected from fluoro andhydroxymethyl. In certain embodiments, R⁴ is selected from fluoro andhydroxymethyl. In certain embodiments, R⁵ is selected from fluoro andhydroxymethyl.

In certain embodiments, the compounds have formula:

or a pharmaceutically acceptable derivative thereof,

wherein R^(3x) is H, azido, substituted or unsubstituted C1-6 alkyl,substituted or unsubstituted C2-6 alkenyl or substituted orunsubstituted C2-6 alkynyl; R^(4x) is H, C1-6 substituted orunsubstituted alkyl, C2-6 substituted or unsubstituted alkenyl or C2-6substituted or unsubstituted alkynyl and other variables are as definedelsewhere herein.

In certain embodiments, the compounds have formula:

or a pharmaceutically acceptable derivative thereof,

wherein R^(3z) is H, C1-6 alkyl, hydroxyl C1-6 alkyl, halo C1-6 alkyl,azido C1-6 alkyl or OH and the other variables are as defined elsewhereherein. In certain embodiments, R^(3z) is hydrogen C1-6 alkyl orhydroxyl C1-6 alkyl. In certain embodiments, R^(3z) is hydrogen orhydroxy methyl. In certain embodiments, R^(3z) is hydroxy methyl.Optionally, the OH groups are protected, for example as an ester. Incertain embodiments, R^(3z) may be in S or R configuration.

In certain embodiments, the compounds have formula:

or a pharmaceutically acceptable derivative thereof.

In certain embodiments, the compounds have formula:

or a pharmaceutically acceptable derivative thereof,

wherein R^(3y) is H, substituted or unsubstituted C1-6 alkyl,substituted or unsubstituted C2-6 alkenyl or substituted orunsubstituted C2-6 alkynyl; or OH and the other variables are as definedelsewhere herein. In certain embodiments, R^(3y) is hydrogen C1-6 alkylor hydroxyl C1-6 alkyl. In certain embodiments, R^(3y) is hydrogen orhydroxy methyl. In certain embodiments, R^(3y) may be in S or Rconfiguration.

In certain embodiments, B is selected from a natural or non naturalpurine or pyrimidine base. In certain embodiments, the base is selectedfrom pyrimidin-1-yl, pyrimidin-3-yl, purin-3-yl, purin-7-yl orpurin-9-yl residue. In certain embodiments, the base is thymin-1-yl,cytosine-1-yl, adenine-9-yl or guanine-9-yl.

a). P^(x) as an Antiviral Phosphonate

In certain embodiments, P^(x) is an antiviral phophonate, including, butnot limited to adefovir, cidofovir, cyclic cidofovir, tenofovir,9-(2-phosphonylmethoxyethyl)guanine (PMEG), 9-(2phosphonyl-methoxyethyl)adenine (PMEA), penciclovir, and9-(3hydroxy-2-phosphonylmethoxypropyl)adenine (HPMPA). In certainembodiments, the phosphonates for use in the compounds provided hereincontain either a phosphonate (—PO₃H₂) or a methylene phosphonate(—CH₂—PO₃H₂) group substituted for the 5′-hydroxyl of the antiviralnucleoside. In other embodiments, P^(x) is a phosphonate derivative ofazidothymidine (AZT). In certain embodiments, P^(x) is cidofovir, cycliccidofovir, tenofovir or 9-(3hydroxy-2-phosphonylmethoxypropyl)adenine(HPMPA). In certain embodiments, P^(x) is cidofovir or cyclic cidofovir.In certain embodiments, P^(x) is9-(3hydroxy-2-phosphonylmethoxypropyl)adenine.

In certain embodiments, P^(x) is an antiviral nucleoside, includingcyclic and acyclic nucleosides, that can be converted to itscorresponding 5′-phosphonate. Such phosphonate analogs typically containeither a phosphonate (—PO₃H₂) or a methylene phosphonate (—CH₂—PO₃H₂)group substituted for the 5′-hydroxyl of the antiviral nucleoside. Someexamples of antiviral phosphonates derived by substituting —PO₃H₂ forthe 5′-hydroxyl are:

3′-azido-3′,5′- dideoxythyinidine-5′- phosphonic acid (AZT phosphonate)

Hakimelahi, G. et al. 3′,5′- dideoxythymidine-2′-ene- 5′-phosphonic acid(d4T phosphonate)

Ibid. 2′,3′5′- trideoxycytidine-5′- phosphonic acid (ddC phosphonate)

Kofoed, T., et al. 9-(3-(phosphono- methoxy)propyl)adenine (Adefovir)

Kim, et al.

Some examples of antiviral phosphonates derived by substituting—CH₂—PO₃H₂ for the 5′-hydroxyl are:

Ganciclovir phosphonate

Huffman, et al. Acyclovir phosphonate

Ibid. Ganciclovir cyclic phosphonate

Smee, et al. 3′-thia-2′,3′- dideoxycytidine-5′- phosphonic acid

Kraus,, et al.

In other embodiment, antiviral nucleotide phosphonates for use in thecompounds provided herein are obtained from antiviral nucleosidesincluding ddA, ddI, ddG, L-FMAU, DXG, DAPD, L-dA, L-dI, L-(d)T, L-dC,L-dG, FTC, penciclovir, and the like.

b). P^(x) as an Anti-Neoplastic

In certain embodiment, P^(x) is an anti-neoplastic selected from avariety of phosphonate-containing nucleotides (or nucleosides which canbe derivatized to their corresponding phosphonates).

In other embodiments, P^(x) is a phosphonate derivative of ananti-neoplastic nucleoside, including, but not limited to cytosinearabinoside, gemcitabine, 5-fluorodeoxyuridine riboside,5-fluorodeoxyuridine deoxyriboside, 2-chlorodeoxyadenosine, fludarabine,1-β-D-arabinofuranosyl-guanine, or pharmaceutically acceptablederivatives thereof.

In certain embodiments, P^(x) is a phosphonate derivative of ananti-neoplastic nucleoside, including, but not limited to2-chloro-deoxyadenosine, 1-β-D-arabinofuranosyl-cytidine (cytarabine,ara-C), fluorouridine, fluorodeoxyuridine (floxuridine), gemcitabine,cladribine, fludarabine, pentostatin (2′-deoxycoformycin),6-mercaptopurine, 6-thioguanine, and substituted or unsubstituted1-β-D-arabinofuranosyl-guanine (ara-G), 1-β-D-arabinofuranosyl-adenosine(ara-A), 1-β-D-arabinofuranosyl-uridine (ara-U).

In certain embodiments P^(x) is 9-(2-phosphonylmethoxyethyl)guanine(PMEG), 9-(2-(phosponomethoxy)ethyl)-2,6-diaminopurine (PMEDAP),9-(2-(phosphonomethoxy)ethyl)adenine (adefovir), and the like.

c). P^(x) as Bisphosphonate

In certain embodiments, P^(x) is a bisphosphonate, such as for example,alendronate, etidronate, tiludronate, ibandronate, Disodium1-hydroxy-3-(1-pyrrolidinyl)-propylidene-1,1-bisphosphonate (EB-1053),pamidronate, olpadronate, amino-olpadronate, clodronate, risedronate,and the like.

In certain embodiments, the phosphonates for use in compounds of formulaI provided herein are bisphosphonate compounds that have the ability toinhibit squalene synthase and to reduce serum cholesterol levels in asubject. Examples of these bisphosphonates are disclosed, for example,in U.S. Pat. Nos. 5,441,946 and 5,563,128, both of which are herebyincorporated by reference in their entirety. In certain embodiments, thebisphosphonate compounds that have the ability to inhibit squalenesynthase are represented by formula:

where: A is O, S, NR, SO, SO₂ or a bond;

B is (CR^(x)R^(x))₁₋₂, O, S, NR^(x), SO, SO₂, R^(x)C═CR^(x), C≡C, O═C ora bond;

Y is:

Z is

W is H,

—COOR_(7′) or SO₃R_(8′)R_(9′); R^(x) is hydrogen or alkyl;

R′ and R″ are independently hydrogen, alkyl, alkoxy, hydroxy, halo,haloalkyl or phenyl; or R′ and R″ together may form a double bond;

each R^(z) is independently hydrogen, alkyl, alkoxy, hydroxy, halo,haloalkyl or phenyl;

R_(3′), R_(4′), R_(5′), R_(7′), R_(7′), R_(8′), and R_(9′) areindependently hydrogen, alkyl, aryl, aralkyl or —CH₂OCOR_(x);

Alk is bi- or tri-carbocycloalkane;

Ar I and Ar II are independently a mono- or di-aryl or heteroaryl;

a and b are independently 0-3;

a+b is 0-4;

d is 0-3;

a+b+d is 1-3;

e is 0-3;

f is 1-6;

m and n are independently 0-2;

x′ is 1-6;

y is 0-2;

x′+y is 3-6; and

its stereoisomers, enantiomers, diastereoisomers and racemic mixtures;or a pharmaceutically acceptable salt thereof.

d). Other Phosphonates for Use in the Alkyl Ester Compounds of Formula I

Several other phosphonate compounds known to those of skill in the artcould be used for preparing the alkyl ester derivatives provided herein.Exemplary phosphonates that can be derivatized as provided are disclosedin the following patents, each of which are hereby incorporated byreference in their entirety: U.S. Pat. No. 3,468,935 (Etidronate), U.S.Pat. No. 4,327,039 (Pamidronate), U.S. Pat. No. 4,705,651 (Alendronate),U.S. Pat. No. 4,870,063 (Bisphosphonic acid derivatives), U.S. Pat. No.4,927,814 (Diphosphonates), U.S. Pat. No. 5,043,437 (Phosphonates ofazidodideoxynucleosides), U.S. Pat. No. 5,047,533 (Acyclic purinephosphonate nucleotide analogs), U.S. Pat. No. 5,142,051(N-Phosphonylmethoxyalkyl derivatives of pyrimidine and purine bases),U.S. Pat. No. 5,183,815 (Bone acting agents), U.S. Pat. No. 5,196,409(Bisphosphonates), U.S. Pat. No. 5,247,085 (Antiviral purine compounds),U.S. Pat. No. 5,300,671 (Gem-diphosphonic acids), U.S. Pat. No.5,300,687 (Trifluoromethylbenzylphosphonates), U.S. Pat. No. 5,312,954(Bis- and tetrakis-phosphonates), U.S. Pat. No. 5,395,826(Guanidinealkyl-1,1-bisphosphonic acid derivatives), U.S. Pat. No.5,428,181 (Bisphosponate derivatives), U.S. Pat. No. 5,442,101(Methylenebisphosphonic acid derivatives), U.S. Pat. No. 5,532,226(Trifluoromethybenzylphosphonates), U.S. Pat. No. 5,656,745 (Nucleotideanalogs), U.S. Pat. No. 5,672,697 (Nucleoside-5′-methylenephosphonates), U.S. Pat. No. 5,717,095 (Nucleotide analogs), U.S. Pat.No. 5,760,013 (Thymidylate analogs), U.S. Pat. No. 5,798,340 (Nucleotideanalogs), U.S. Pat. No. 5,840,716 (Phosphonate nucleotide compounds),U.S. Pat. No. 5,856,314 (Thio-substituted, nitrogen-containing,heterocyclic phosphonate compounds), U.S. Pat. No. 5,885,973(olpadronate), U.S. Pat. No. 5,886,179 (Nucleotide analogs), U.S. Pat.No. 5,877,166 (Enantiomerically pure 2-aminopurine phosphonatenucleotide analogs), U.S. Pat. No. 5,922,695 (Antiviral phosphonomethoxynucleotide analogs), U.S. Pat. No. 5,922,696 (Ethylenic and allenicphosphonate derivatives of purines), U.S. Pat. No. 5,977,089 (Antiviralphosphonomethoxy nucleotide analogs), U.S. Pat. No. 6,043,230 (Antiviralphosphonomethoxy nucleotide analogs), U.S. Pat. No. 6,069,249 (Antiviralphosphonomethoxy nucleotide analogs); U.S. Pat. Nos. 6,057,305;6,653,296; Belgium Patent No. 672205 (Clodronate); European Patent No.753523 (Amino-substituted bisphosphonic acids); European PatentApplication 186405 (geminal diphosphonates); and the like. In addition,the compounds listed in the following publications can be derivatizedaccording to the invention to improve their pharmacologic activity, orto increase their oral absorption; each of which are hereby incorporatedby reference in their entirety: J. Med. Chem., 2002, 45:1918-1929; J.Med. Chem., 2003, 46:5064-5073; Antimicrob. Agents Chemotherapy, 2002,46:2185-2193. One of skill in the art would be able to selectappropriate phosphonate compounds for use herein.

The following U.S. patents describe other nucleotide phosphonateanalogs: U.S. Pat. No. 5,672,697 (Nucleoside-5′-methylene phosphonates),U.S. Pat. No. 5,922,695 (Antiviral phosphonomethoxy nucleotide analogs),U.S. Pat. No. 5,977,089 (Antiviral phosphonomethoxy nucleotide analogs),U.S. Pat. No. 6,043,230 (Antiviral phosphonomethoxy nucleotide analogs),U.S. Pat. No. 6,069,249, U.S. Pat. No. 5,792,756 (Prodrugs ofPhosphonates), U.S. Pat. No. 5,869,468 (Treatment of Conditions ofAbnormally Increased Intraocular Pressure by Administration ofPhosphonylmethoxyalkyl Nucleoside Analogs and Related NucleosideAnalogs), U.S. Pat. No. 5,854,228 (Antiviral PhosphonomethoxyalkylenePurine and Pyrimidine Derivatives) and U.S. Pat. No. 5,663,159 (Prodrugsof Phosphonates). Also describing nucleotide phosphonates is EP 0 269947 (Antiviral phosphonomethoxyalkylene purine and pyrimidineDerivatives) and EP 0 481 214 (Prodrugs of Phosphonates).

The phosphonates compounds known in the art can be converted in theiralkyl ester as described herein. One of skill in the art can easilyselect an appropriate phosphonate compound for esterification. Somephosphonate compounds that can be esterified are described herein. Theappropriate phosphonate for esterification as described herein containsa free —OH group that can be derivatized into its alkyl ester. Incertain embodiments, the phosphonate compounds have formula 1:

in which R_(1C)-R_(11C) can be the same or different and stand forhydrogen, a straight or branched aliphatic or alicyclic C₁-C₁₀hydrocarbon radical, an aryl or an aryl-C₁-C₄-alkyl radical; n is zeroor one, and m is zero, one or two, or R_(2c) and R_(4c) when takentogether can form a saturated aliphatic 5-, 6- or 7-membered ring whichmay be substituted with one or more C₁-C₄-alkyl radicals.

In certain embodiments, the phosphonates for use in the alkyl estercompounds provided herein are represented by formula 2:

wherein B represents a substituted or unsubstituted purine base,including, but not limited to adenine or guanine and their halogenatedderivatives, R_(1d) is selected from H, methyl, hydroxymethyl,halomethyl, azidomethyl, and cyano; R_(2d) is selected from H, methyl,hydroxymethyl, halomethyl, azidomethyl, cyano, and OH; also when R_(2d)is OH, the carbon to which it is attached may be oxidized so that the Hthere shown and R_(2d) together may be ═O; and n is an integer of 0-5.

In other embodiments, the phosphonate compounds for use herein arerepresented by formula 3:

wherein R_(e) is a hydrogen atom or a hydroxymethyl group and B₁ is apyrimidin-1-yl, pyrimidin-3-yl, purin-3-yl, purin-7-yl or purin-9-ylresidue but not an adenin-9-yl residue, and the salts thereof withalkali metals, ammonia or amines.

In other embodiments, the phosphonate compounds for use herein arerepresented by formula 4:A2-B2-C2

wherein:

A2 is a residue of a hydroxyl containing steroidal hormone possessinghuman bone resorption antagonist activity or bone formation stimulatoryactivity;

C2 is a residue of an amino or hydroxy alkyl-1,1-bisphosphonate,possessing human bone affinity; and

B2 is a covalent linkage, connecting A2 through the hydroxyl moiety andC2 through the respective amino or hydroxyl moiety, which linkage canhydrolyze in the human body in the vicinity of bone to release steroidalhormone A2, and pharmaceutically acceptable salts or esters thereof.

In other embodiments, the phosphonate compounds for use herein arerepresented by formula 5:

wherein m₁ and l₁ are independently 1 or 2;R_(1f) represents hydrogen, a lower alkyl group, or an alkali metalcation;R_(2f) represents hydrogen, a lower alkyl group or an alkali metalcation;Y represents ═O or ═N—OH, or —OH; andX represents —(CH₂)n_(f)-, a branched alkylene group, or a branched orstraight alkenylene or alkynylene chain optionally substituted by one ormore oxygen or nitrogen atoms, wherein n_(f) is an integer from 3 to 24;with the provision that when l₁=m₁=1, Y is ═O, and R_(1f) and R_(2f)denote a lower alkyl group, n_(f) represents an integer from 9 to 20;and with the provision that when l₁=m₁=2, Y is —OH and R_(1f) and R_(2f)are methyl groups, n_(f) represents an integer from 9-24; and with thefurther provision that when l₁=m₁=2, Y is —OH and R_(1f) and R_(2f) arehydrogen, n_(f) represents 3, 5 or an integer from 7 to 24; orX is —(CH₂)_(P)—(OCH₂CH₂)_(q)—O—(CH₂)p′-, wherein p and p′ areindependently integers from 1 to 5 and q is an integer from 1 to 6; orX is —(CH₂)t-O—(CH₂)s-O—(CH₂)t′- wherein t and t′ are independentlyintegers from 1 to 6 and s is an integer from 2 to 12; orX is—B—A—B

Wherein B represents a branched group or straight alkylene, or analkenylene or alkynylene chain optionally substituted by one or moreoxygen or nitrogen atoms, and A represents an aromatic group such asphenylene, naphthalenediyl, thiophenediyl or furandiyl. In certainembodiments, Y represents ═O or ═N—OH.

In other embodiments, the phosphonate compounds for use herein arerepresented by formula 6:

or a pharmaceutically acceptable salt thereof:wherein R_(1g) is hydroxy, amino, chloro or OR_(7g) wherein R_(7g) isC1-6 alkyl, phenyl or phenyl C1-2 alkyl either of which phenyl moietiesmay be substituted by one or two substituents selected from halo, C1-4alkyl or C1-4 alkoxy;R_(2g) is amino or, when R_(1g) is hydroxy or amino, R₂₉ may also behydrogen;R_(3g) is hydrogen, hydroxymethyl or acyloxymethyl;R_(4g) is a group of formula:

wherein

R_(5g) and R_(6g) are independently selected from hydrogen, C1-6 alkyland optionally substituted phenyl; or R_(3g) and R_(4g) together are:

wherein R_(6g) is as defined above.

In other embodiments, the phosphonate compounds for use herein arerepresented by formula 7:

wherein:

-   -   R_(1h) and R_(2h), which can be the same or different, are        hydrogen or C1-C4 alkyl;

(A) is hydrogen, halogen, hydroxy, straight or branched C1-C12 alkyl;

(B) is a covalent bond, a straight or branched C1-C8 alkylene or,together with the adjacent nitrogen atom, a group of formula

therein the groups —N(R_(3h))— and —(CH₂)n_(h)- may be in the 1,1; 1,2;1,3 or 1,4 position of the ring; an ortho, meta or para-substitutedaralkylene of formula

an alkylene chain containing at least one hetero-atom of formula—[CH(CH3)]p-(CH2)nl-X—(CH2)n_(h)-; m_(h) is the integer 5 or 6; n_(h)and nl are an integer from 1 to 5; p is zero or 1 and X is O, S, —N—CH₃or the ureido group —NH—CO—NH—; R_(3h) is hydrogen, straight or branchedC1-C9 alkyl, C3-C6 cycloalkyl, benzyl, phenyl or p-methoxybenzyl;

(C) is straight or branched C1-C5 alkyl, phenylene, an aralkylene chainof formula

in which n_(h) is as above defined; R_(4h) is hydrogen, straight orbranched C1-C4 alkyl, or it is a group of formula

in which R_(7h) and R_(8h), which are the same or different, arehydrogen, straight or branched C1-C6 alkyl, phenyl, benzyl,p-methoxybenzyl, or one of R_(7h) and R_(8h) is as above defined and theother one is a group of formula R_(9h)—C(O)—, in which R_(9h) ishydrogen, straight or branched C1-C4 alkyl, phenyl, benzyl,p-methoxyphenyl, straight or branched C1-C4 alkoxy, halo-C1-C4-alkoxy;

R_(5h) and R_(6h) are haloethyl (2-chloroethyl, 2-bromoethyl,2-iodoethyl) or R_(5h) and R_(6h), together with the nitrogen atom towhich they are bound, are a 1-aziridinyl residue of formula

In other embodiments, the phosphonate compounds for use herein arerepresented by formula 8:

wherein R_(1i) and R_(2i) are the same or different and are selectedfrom any of hydrogen, C1-C8 alkyl, C1-C8 alkenyl, hydroxyalkyl whereinthe alkyl portion is C1-C8, alkoxyalkyl wherein the alkyl portion isC1-C8, aralkyl wherein the alkyl portion is C1-C8, such as benzyl,phenylethyl, phenylpropyl, aryl such as phenyl or aminoalkyl, theaminoalkyl includes substituents of the formula —(CH₂)n-NR_(7i)R_(8i),wherein n=2-6 and R_(7i) and R_(8i) are the same or different and areselected from any of H, C1-C4 alkyl, aralkyl wherein the alkyl portionin C1-C4 or R_(7i) and R_(8i) may be taken together to form aheterocyclic ring having from 5 to 7 ring atoms containing one or moreheteroatoms such as N, O and S, examples of suitable ring systemsinclude piperidino, morpholino, tetrahydroquinolinyl,tetrahydroisoquinolinyl, thiomorpholino and piperazino substituted atthe N-4 position by R_(9i), wherein R_(9i) is selected from any one ofC1-C4 alkyl, or aralkyl wherein the alkyl portion is C1-C4 alkyl orphenyl;

R_(3i) and R_(5i) may each be either H or CF₃, with the proviso thatonly one of R_(3i) or R_(5i) may be CF₃ at the same time;

R_(4i) and R_(6i) may each be H or CF₃, with the proviso that if eitheror both of R_(4i) and R_(6i) are CF3, neither R_(3i) nor R_(5i) may beCF₃ and with the further proviso that R_(3i)-R_(6i) may not each be H atthe same time.

In other embodiments, the phosphonate compounds for use herein arerepresented by tautomeric formula 9a, 9b or 9c

and/or a pharmaceutically acceptable salt of the compound of the formula9a, 9b or 9c;

R^(J) has the following meaning therein:

I) a radical of the formula

wherein R^(11J) or R^(12J) has the following meaning: a)R^(13J)—S(O)n_(J)—

or b)

where R^(13J) is 1) (C1-C6)-alkyl, 2) (C5-C7)-cycloalkyl, 3)cyclopentylmethyl, 4) cyclohexylmethyl, 5) phenyl, 6) phenyl substitutedonce to three times by fluorine atom, chlorine atom, methyl or methoxy,where n_(J) is the integer zero, 1 or 2, where R^(14J) and R^(15J) areidentical or different and have, independently of one another, thefollowing meaning: 1) hydrogen atom, 2) (C1-C6)-alkyl, 3) phenyl, 4)phenyl substituted once or twice by fluorine atom, chlorine atom, methylor methoxy, 5) —(CH2)m_(J)-phenyl where m_(J) is an integer from 1 to 4,or 6) —(CH2)m_(J)-phenyl where m is an integer from 1 to 4 and thephenyl radical is substituted once or twice by the radicals indicated in4, 7) R^(14J) and R^(15J) together form a straight-chain or branchedchain of 4 to 7 carbon atoms, the chain can additionally be interruptedby O, S or NR where R is hydrogen or methyl, or 8) R^(14J) and R^(15J)form together with the nitrogen atom to which they bonded adihydroindole, tetrahydroquinoline or tetrahydroisoquinoline ring, andthe other substituent R^(11J) or R^(12J) in each case means a) hydrogenatom, b) halogen atom, such as fluorine, chlorine, bromine or iodineatom, c) (C1-C4)-alkyl, d) (C1-C4)-alkoxy, e) phenoxy, f) phenoxysubstituted once to three times by fluorine, chlorine, methyl ormethoxy, g) R^(13J)—S(O)n_(J), where n_(J) is the integer zero, 1 or 2and R^(13J) has the abovementioned meaning, or h)

where R^(14J) and R^(15J) have the abovementioned meaning, or

II) a radical of the formula

wherein R^(21J), R^(22J) or R^(23J) has the following meaning: a)hydrogen atom,b) halogen atom, such as fluorine, chlorine, bromine or iodine atom, orc) (C1-C12)-alkyl,where one of the substituents R^(21J), R^(22J) or R^(23J) can also mean1) N₃, 2) CN, 3) OH, 4) (C1-C10)-alkoxy, 5) R^(24J)—C_(nJ)H._(2nJ)—O_(mJ), wherem_(J) is the number zero or 1, nj is the number zero, 1, 2 or 3,R^(24J) is 1) C_(p)F_(2p)+1 where p is the number 1, 2 or 3, as long asnj is the number zero or 1, 2) (C3-C12)-cycloalkyl, 3) phenyl, 4)pyridyl, 5) quinolyl or 6) isoquinolyl, where the ring system in theradicals 3) to 6) is unsubstituted or substituted by a radical from thegroup fluorine atom, chlorine atom, CF3, methyl, methoxy orNR^(25J)R^(26J) where R^(25J) and R^(26J) are identical or different andhave, independently of one another, the meaning hydrogen atom or(C1-C.4)-alkyl, or

III) a radical of the formula

wherein R^(31J), R^(32J), R^(33J) or R^(34J) independently has thefollowing meaning: a) hydrogen atom, b) halogen atom, such as fluorine,chlorine, bromine or iodine atom, or c) —CN, d) —NO₂, e) —N₃, f)—(C1-C6)-alkyl, straight-chain or branched or g) R^(35J)—CnH_(2n)—Z—,where n is the number zero, 1, 2, 3, 4, 5 or 6, and the alkylene chain—CnH2n- is straight-chain or branched, and one carbon atom can bereplaced by an oxygen, sulfur or nitrogen atom, R^(35J) is 1) hydrogenatom, 2) (C3-C6)-alkenyl, 3) (C5-C8)-cycloalkyl, 4) (C5-C8)-cycloalkyl,substituted by a hydroxyl group, or one methylene group is replaced byan oxygen, sulfur or nitrogen atom, or 5) phenyl, unsubstituted orsubstituted by 1 to 3 radicals from the group halogen atom such asfluorine, chlorine, bromine or iodine atom, CF₃; CH₃—S(O)x, where x isthe number zero, 1 or 2; R^(36J)—Wy where R^(36J) is hydrogen atom,methyl or ethyl, W is oxygen atom, NH or NCH₃, and y is zero or 1;C_(m)F_(2m+1), where m is the number 1, 2 or 3; pyridyl; quinolyl orisoquinolyl,

Y is CH₂ or NH;

Z is

1) —CO—, 2) —CH₂—, 3) —[CH(OH)]q-, where q is the number 1, 2 or 3, 4)—[C(CH₃)(OH)]q-, where q is the number 1, 2 or 3, 5) —O—, 6) —NH—, 7)—N(CH₃)—, 8) —S(O)x-, where x is zero, 1 or 2, 9) —SO2-NH— or 10)

X has the following meaning a) N or b) C—R^(37J), where R^(37J) ishydrogen atom, (C1-C4)-alkyl or (C2-C4)-alkenyl,

Y has the following meaning

a) NH, b) —N—(C2-C6)-alkyl, c) —N—(C2-C4)-alkenyl or d)R^(35J)—CnH2n-Z—, where R^(35J), n and Z are defined as above, R^(5J),R^(6J), R^(7J) and R^(8J), are identical or different and have,independently of one another, the following meaning a) hydrogen atom, b)(C1-C5)-alkyl, straight-chain or branched, or c) phenyl.

In other embodiments, the phosphonate compounds for use herein arerepresented by formula 10:

wherein Ak—O— denotes a residue of a compound having an estrogenicactivity; R_(1k) independently denotes H or a C1-C6 alkyl group; Xkdenotes a single bond, a C1-C10 alkylene group or a group of theformula:

wherein R_(2k) denotes H or a C1-C5 alkyl group; Zk denotes a nitrogroup or a halogen; n′ is an integer of 3 to 12; k is an integer of 1 to5; L is an integer of 0 to 5; and q is an integer of 1 to 3, andphysiologically acceptable salts thereof.

In other embodiments, the phosphonate compounds for use herein are5′-phosphonates of 3′-azido-2′,3′-dideoxynucleosides that arerepresented by formula 11:

and B¹ is thymin-1-yl, cytosine-1-yl, adenine-9-yl or guanine-9-yl.

In other embodiments, the phosphonate compounds for use herein aremethylenebisphosphonic acid derivatives represented by formula 10:

in which W₁, W₂, W₃ and W₄ are independently the group OR_(1m) or thegroup NR_(2m) R_(3m) wherein R_(1m), R_(2m) and R_(3m) independently arehydrogen or straight or branched, optionally unsaturated C1-C22-alkyl,optionally substituted, optionally unsaturated C3-C10-cycloalkyl, aryl,aralkyl or silyl SiR_(3m), or the groups R_(2m) and R_(3m) form togetherwith the adjacent nitrogen atom a 3 to 10-membered saturated, partlysaturated or aromatic heterocyclic ring, wherein in addition to thenitrogen atom, there may be one or two heteroatoms from the group N, Oand S, provided that in the formula I at least one of the groups W₁, W₂,W₃ and W₄ is hydroxy and at least one of the groups W₁, W₂, W₃ and W₄ isthe amino group NR_(2m)R_(3m), R_(3′) is lower alkyl

Q¹ and Q² are independently hydrogen, fluorine, chlorine, bromine oriodine, including the stereoisomers, such as the geometrical isomers andthe optically active isomers, of the compounds, as well as thepharmacologically acceptable salts of the compounds.

In other embodiments, the phosphonate compounds for use herein arerepresented by formula 11:

wherein B_(n) is independently a purine or pyrimidine base or modifiedform, each R_(1n) is independently hydrogen, hydroxyl, fluorine ormethyl ester; each Y_(n) is independently OR_(2n), N(R_(2n))₂ or SR_(2n)wherein, each R_(2n) is independently hydrogen or alkyl (1-12 C); X isselected from oxygen and sulfur.

In other embodiments, the phosphonate compounds for use herein arerepresented by formula 12:

where B₃ is a protected or unprotected cytosin-1-yl.

In other embodiments, the phosphonate compounds for use herein arerepresented by formula 13:

where B⁰ is a protected or unprotected heterocyclic base;

R is hydrogen (H), alkyl, O-alkyl, —CHO, —C(O)OR^(2o), —C(O)R^(2o),—C(O)N(R^(3o))₂ or —S(O)₂ N(R^(3o))₂;

each R^(o) is independently hydrogen, cyano (CN), nitro (NO₂), halogen,alkyl, O-alkyl, —C(O)OR^(3o), —C(O)R^(3o), —S(O)₂ OH, —N(R^(3o))₂, —CHOor —OH; and

each R^(2o) and each R^(3o) are independently hydrogen, alkyl, phenyl,alkyl substituted phenyl, —CH₂C6H₅ or —CH₂CH₂C6H₅.

In other embodiments, the phosphonate compounds for use herein arerepresented by formula 14:

wherein: R^(p) is selected from the group consisting of

in which R^(1p) and R^(2p) are independently selected from the groupconsisting of hydrogen and lower alkyl, R^(3p) is lower alkyl or—(CH₂)n_(p)—C₆H₅, and n_(p) is an integer in the range of 0 to 6inclusive;

R′ is selected from the group consisting of hydrogen, hydroxyl,carboxyl, alkoxy, amino and halogen; and

R″ is hydrogen or a halogen substituent, or a pharmaceuticallyacceptable salt or ester thereof.

In other embodiments, the phosphonate compounds for use herein arerepresented by formula 15:

wherein Zq is independently —OC(R^(2q))₂OC(O)Xq(R^(q))_(a), an ester, anamidate or —H, but at least one Zq is —OC(R^(2q))₂OC(O)Xq(R^(q))_(a);

A^(q) is the residue of an antiviral phosphonomethoxy nucleotide analog;

Xq is N or O;

R^(2q) independently is —H, C1-C12 alkyl, C5-C12 aryl, C2-C12 alkenyl,C2-C12 alkynyl, C7-C12 alkenylaryl, C7-C12 alkynylaryl, or C6-C1-2alkaryl, any one of which is unsubstituted or is substituted with 1 or 2halo, cyano, azido, nitro or —OR^(3q) in which R^(3q) is C1-C1-2 alkyl,C2-C12 alkenyl, C2-C1 alkynyl or C5-C12 aryl;

R^(q) independently is —H, C1-C12 alkyl, C5-C12 aryl, C2-C12 alkenyl,C2-C12 alkynyl, C7-C12 alkyenylaryl, C7-C12 alkynylaryl, or C6-C12alkaryl, any one of which is unsubstituted or is substituted with 1 or 2halo, cyano, azido, nitro, —N(R⁴)₂ or —OR where R_(4q) independently is—H or C1-C8 alkyl, provided that at least one R is not H; and

a is 1 when Xq is O, or 1 or 2 when Xq is N;

with the proviso that when a is 2 and Xq is N, (a) two N-linked R^(q)groups can be taken together to form a carbocycle or oxygen-containingheterocycle, (b) one N-linked R^(q) additionally can be —OR^(3q) or (c)both N-linked R^(q) groups can be —H; and the salts, hydrates, tautomersand solvates thereof.

e). Exemplary Esterified Phosphonate Compounds

In certain embodiments, the compounds herein are alkyl esters ofcidofovir, cyclic cidofovir or HPMPA. In certain embodiments, the alkylesters of CDV possess 16-22 carbon atoms. In certain embodiments, thealkyl esters of CDV possess 16, 17, 18, 19, 20, 21 or 22 carbon atoms.In certain embodiments, the alkyl esters of CDV possess 18, 19 or 20carbon atoms In certain embodiments, the alkyl esters of cidofovir areselected from octyl cidofovir, dodecyl cidofovir, hexadecyl cidofovir,eicosyl cidofovir, docosyl cidofovir and tetracosyl cidofovir. In otherembodiment, the alkyl esters of cidofovir are selected from eicosylcidofovir, docosyl cidofovir and tetracosyl cidofovir. In certainembodiments, the alkyl esters of cidofovir are selected from octylcyclic cidofovir, dodecyl cyclic cidofovir, hexadecyl cyclic cidofovir,eicosyl cyclic cidofovir, docosyl cyclic cidofovir and cyclic cyclictetracosyl cidofovir. In other embodiment, the alkyl esters of cidofovirare selected from cyclic eicosyl cidofovir, docosyl cyclic cidofovir andtetracosyl cyclic cidofovir. In other embodiment, the alkyl esters ofHPMPA is eicosyl-(S)-HPMPA. In certain embodiments, the compoundsprovided herein are 7-methyl-octyl, 8-methyl-nonyl, 9-methyl-decyl,10-methyl-undecyl, 11-methyl-dodecyl, 12-methyl-tridecyl,13-methyl-tetradecyl, 14-methyl-pentadecyl, 15-methyl-hexadecyl,16-methyl-heptadecyl, 17-methyl-octadecyl, 18-methyl-nonadecyl,19-methyl-eicosyl, 20-methyl-heneicosyl, 21-methyl-docosyl,22-methyl-tricosyl, 7-fluoro-octyl, 8-fluoro-nonyl, 9-fluoro-decyl,10-fluoro-undecyl, 11-fluoro-dodecyl, 12-fluoro-tridecyl,13-fluoro-tetradecyl, 14-fluoro-pentadecyl, 15-fluoro-hexadecyl,16-fluoro-heptadecyl, 17-fluoro-octadecyl, 18-fluoro-nonadecyl,19-fluoro-eicosyl, 20-fluoro-heneicosyl, 21-fluoro-docosyl or22-fluoro-tricosyl ester of cidofovir or cyclic cidofovir.

In one embodiment, a derivative of cyclic cidofovir is provided whichincludes an alkyl ester, wherein the alkyl is a C18-22 straight,branched, or cyclic alkyl or alkenyl having 1 to 6 double bonds.

In certain embodiments, the compounds provided herein possess one ormore chiral centers, e.g. in the sugar moieties, and may thus exist inoptically active forms. Likewise, when the compounds contain an alkenylgroup or an unsaturated alkyl or acyl moiety there exists thepossibility of cis- and trans-isomeric forms of the compounds.Additional asymmetric carbon atoms can be present in a substituent groupsuch as an alkyl group. The R- and S-isomers and mixtures thereof,including racemic mixtures as well as mixtures of cis- and trans-isomersare contemplated herein. All such isomers as well as mixtures thereofare intended to be included within the scope of the compounds providedherein. If a particular stereoisomer is desired, it can be prepared bymethods well known in the art by using stereospecific reactions withstarting materials that contain the asymmetric centers and are alreadyresolved or, alternatively, by methods that lead to mixtures of thestereoisomers and resolution by known methods.

C. Preparation of the Compounds

The alkyl ester derivatives of phosphonates for use in the compositionsand methods provided herein can be prepared by alkyl esterification ofvarious phosphonate compounds. Exemplary methods for esterification aredescribed herein but other esterification methods well known in the artcan be used to prepare the phosphonate alkyl esters provided herein.

Scheme 1 demonstrates a general procedure for esterification of aphosphonate compound by reacting it with a suitable bromoalkane.

An appropriate 1-bromoalkane (1.2 eq) is added to a suspension of aphosphonate compound in a suitable solvent such asN,N-dicyclohexyl-4-morpholino-carboxamidine (1.1 eq) andN,N-dimethylformamide (10 ml/mmol). The mixture is heated and stirredmagnetically overnight. The solvent is then evaporated under reducedpressure, and the residue is purified. The purification can be carriedout by methods known to those of skill in the art, including, but notlimited to flash column chromatography to isolate the phosphonate alkylesters.

Scheme 2 demonstrates an alternate general procedure for esterificationof a phosphonate compound using Mitsunobu reaction.

A phosphonate compound (1 eq), an appropriate alkanol (2 eq) andtriphenyl phosphine (2 eq) are mixed in anhydrous DMF. The mixture isstirred under nitrogen. Diisopropyl azadicarboxylate (2 eq) is thenadded in three portions over 15 min before the mixture is allowed tostir overnight. The solvent is then evaporated under vacuum, and theresidue is purified by methods known in the art.

The alkyl esters of cyclic cidofovir analogs provided herein can beconverted to their corresponding acyclic analogs by procedures known inthe art. For example, an alkyl-cCDV analog is suspended in 2 M NaOH (25ml/mmol). The suspension is heated to 80° C. and stirred for 1 h. Afterhydrolysis, the solution is cooled to 25° C. and acidified with glacialacetic acid (pH approximately 5). The resulting precipitate is collectedby vacuum filtration and dried under vacuum. The crude product ispurified by known method.

Other methods known to those of skill in the art can be similarly usedfor the preparation of alkyl ester compounds provided herein.

D. Formulation of Pharmaceutical Compositions

The pharmaceutical compositions provided herein contain therapeuticallyeffective amounts of one or more of the compounds provided herein thatare useful in the prevention, treatment, or amelioration of one or moreof the symptoms of diseases or disorders associated with viralinfections, inappropriate cell proliferation or bone metabolism and apharmaceutically acceptable carrier. Pharmaceutical carriers suitablefor administration of the compounds provided herein include any suchcarriers known to those skilled in the art to be suitable for theparticular mode of administration.

In addition, the compounds may be formulated as the solepharmaceutically active ingredient in the composition or may be combinedwith other active ingredients.

The compositions contain one or more compounds provided herein. Thecompounds are, in one embodiment, formulated into suitablepharmaceutical preparations such as solutions, suspensions, tablets,dispersible tablets, pills, capsules, powders, sustained releaseformulations or elixirs, for oral administration or in sterile solutionsor suspensions for parenteral administration, as well as transdermalpatch preparation and dry powder inhalers. In one embodiment, thecompounds described above are formulated into pharmaceuticalcompositions using techniques and procedures well known in the art (see,e.g., Ansel Introduction to Pharmaceutical Dosage Forms, Fourth Edition1985, 126).

In the compositions, effective concentrations of one or more compoundsor pharmaceutically acceptable derivatives thereof is (are) mixed with asuitable pharmaceutical carrier. The compounds may be derivatized as thecorresponding salts, esters, enol ethers or esters, acetals, ketals,orthoesters, hemiacetals, hemiketals, acids, bases, solvates, hydratesor prodrugs prior to formulation, as described above. The concentrationsof the compounds in the compositions are effective for delivery of anamount, upon administration, that treats, prevents, or ameliorates oneor more of the symptoms of diseases or disorders associated withassociated with viral infections, inappropriate cell proliferation orbone metabolism. In one embodiment, the compositions are formulated forsingle dosage administration. To formulate a composition, the weightfraction of compound is dissolved, suspended, dispersed or otherwisemixed in a selected carrier at an effective concentration such that thetreated condition is relieved, prevented, or one or more symptoms areameliorated.

The active compound is included in the pharmaceutically acceptablecarrier in an amount sufficient to exert a therapeutically useful effectin the absence of undesirable side effects on the patient treated. Thetherapeutically effective concentration may be determined empirically bytesting the compounds in in vitro and in vivo systems well known tothose of skill in the art and then extrapolated therefrom for dosagesfor humans.

The concentration of active compound in the pharmaceutical compositionwill depend on absorption, inactivation and excretion rates of theactive compound, the physicochemical characteristics of the compound,the dosage schedule, and amount administered as well as other factorsknown to those of skill in the art. For example, the amount that isdelivered is sufficient to ameliorate one or more of the symptoms ofdiseases or disorders associated with viral infections, inappropriatecell proliferation or bone metabolism, as described herein.

In one embodiment, a therapeutically effective dosage should produce aserum concentration of active ingredient of from about 0.1 ng/ml toabout 50-100 μg/ml. The pharmaceutical compositions, in anotherembodiment, should provide a dosage of from about 0.001 mg to about 2000mg of compound per kilogram of body weight per day. Pharmaceuticaldosage unit forms are prepared to provide from about 0.01 mg, 0.1 mg or1 mg to about 500 mg, 1000 mg or 2000 mg, and in one embodiment fromabout 10 mg to about 500 mg of the active ingredient or a combination ofessential ingredients per dosage unit form.

The active ingredient may be administered at once, or may be dividedinto a number of smaller doses to be administered at intervals of time.It is understood that the precise dosage and duration of treatment is afunction of the disease being treated and may be determined empiricallyusing known testing protocols or by extrapolation from in vivo or invitro test data. It is to be noted that concentrations and dosage valuesmay also vary with the severity of the condition to be alleviated. It isto be further understood that for any particular subject, specificdosage regimens should be adjusted over time according to the individualneed and the professional judgment of the person administering orsupervising the administration of the compositions, and that theconcentration ranges set forth herein are exemplary only and are notintended to limit the scope or practice of the claimed compositions.

In instances in which the compounds exhibit insufficient solubility,methods for solubilizing compounds may be used. Such methods are knownto those of skill in this art, and include, but are not limited to,using cosolvents, such as dimethylsulfoxide (DMSO), using surfactants,such as TWEEN®, or dissolution in aqueous sodium bicarbonate.Derivatives of the compounds, such as prodrugs of the compounds may alsobe used in formulating effective pharmaceutical compositions.

Upon mixing or addition of the compound(s), the resulting mixture may bea solution, suspension, emulsion or the like. The form of the resultingmixture depends upon a number of factors, including the intended mode ofadministration and the solubility of the compound in the selectedcarrier or vehicle. The effective concentration is sufficient forameliorating the symptoms of the disease, disorder or condition treatedand may be empirically determined.

The pharmaceutical compositions are provided for administration tohumans and animals in unit dosage forms, such as tablets, capsules,pills, powders, granules, sterile parenteral solutions or suspensions,and oral solutions or suspensions, and oil-water emulsions containingsuitable quantities of the compounds or pharmaceutically acceptablederivatives thereof. The pharmaceutically therapeutically activecompounds and derivatives thereof are, in one embodiment, formulated andadministered in unit-dosage forms or multiple-dosage forms. Unit-doseforms as used herein refers to physically discrete units suitable forhuman and animal subjects and packaged individually as is known in theart. Each unit-dose contains a predetermined quantity of thetherapeutically active compound sufficient to produce the desiredtherapeutic effect, in association with the required pharmaceuticalcarrier, vehicle or diluent. Examples of unit-dose forms includeampoules and syringes and individually packaged tablets or capsules.Unit-dose forms may be administered in fractions or multiples thereof. Amultiple-dose form is a plurality of identical unit-dosage formspackaged in a single container to be administered in segregatedunit-dose form. Examples of multiple-dose forms include vials, bottlesof tablets or capsules or bottles of pints or gallons. Hence, multipledose form is a multiple of unit-doses which are not segregated inpackaging.

Liquid pharmaceutically administrable compositions can, for example, beprepared by dissolving, dispersing, or otherwise mixing an activecompound as defined above and optional pharmaceutical adjuvants in acarrier, such as, for example, water, saline, aqueous dextrose,glycerol, glycols, ethanol, and the like, to thereby form a solution orsuspension. If desired, the pharmaceutical composition to beadministered may also contain minor amounts of nontoxic auxiliarysubstances such as wetting agents, emulsifying agents, solubilizingagents, pH buffering agents and the like, for example, acetate, sodiumcitrate, cyclodextrine derivatives, sorbitan monolaurate,triethanolamine sodium acetate, triethanolamine oleate, and other suchagents.

Actual methods of preparing such dosage forms are known, or will beapparent, to those skilled in this art; for example, see Remington'sPharmaceutical Sciences, Mack Publishing Company, Easton, Pa., 15thEdition, 1975.

Dosage forms or compositions containing active ingredient in the rangeof 0.005% to 100% with the balance made up from non-toxic carrier may beprepared. Methods for preparation of these compositions are known tothose skilled in the art. The contemplated compositions may contain0.001%-100% active ingredient, in one embodiment 0.1-95%, in anotherembodiment 75-85%.

1. Compositions for Oral Administration

Oral pharmaceutical dosage forms are either solid, gel or liquid. Thesolid dosage forms are tablets, capsules, granules, and bulk powders.Types of oral tablets include compressed, chewable lozenges and tabletswhich may be enteric-coated, sugar-coated or film-coated. Capsules maybe hard or soft gelatin capsules, while granules and powders may beprovided in non-effervescent or effervescent form with the combinationof other ingredients known to those skilled in the art.

a. Solid Compositions for Oral Administration

In certain embodiments, the formulations are solid dosage forms, in oneembodiment, capsules or tablets. The tablets, pills, capsules, trochesand the like can contain one or more of the following ingredients, orcompounds of a similar nature: a binder; a lubricant; a diluent; aglidant; a disintegrating agent; a coloring agent; a sweetening agent; aflavoring agent; a wetting agent; an emetic coating; and a film coating.Examples of binders include microcrystalline cellulose, gum tragacanth,glucose solution, acacia mucilage, gelatin solution, molasses,polyinylpyrrolidine, povidone, crospovidones, sucrose and starch paste.Lubricants include talc, starch, magnesium or calcium stearate,lycopodium and stearic acid. Diluents include, for example, lactose,sucrose, starch, kaolin, salt, mannitol and dicalcium phosphate.Glidants include, but are not limited to, colloidal silicon dioxide.Disintegrating agents include crosscarmellose sodium, sodium starchglycolate, alginic acid, corn starch, potato starch, bentonite,methylcellulose, agar and carboxymethylcellulose. Coloring agentsinclude, for example, any of the approved certified water soluble FD andC dyes, mixtures thereof; and water insoluble FD and C dyes suspended onalumina hydrate. Sweetening agents include sucrose, lactose, mannitoland artificial sweetening agents such as saccharin, and any number ofspray dried flavors. Flavoring agents include natural flavors extractedfrom plants such as fruits and synthetic blends of compounds whichproduce a pleasant sensation, such as, but not limited to peppermint andmethyl salicylate. Wetting agents include propylene glycol monostearate,sorbitan monooleate, diethylene glycol monolaurate and polyoxyethylenelaural ether. Emetic-coatings include fatty acids, fats, waxes, shellac,ammoniated shellac and cellulose acetate phthalates. Film coatingsinclude hydroxyethylcellulose, sodium carboxymethylcellulose,polyethylene glycol 4000 and cellulose acetate phthalate.

The compound, or pharmaceutically acceptable derivative thereof, couldbe provided in a composition that protects it from the acidicenvironment of the stomach. For example, the composition can beformulated in an enteric coating that maintains its integrity in thestomach and releases the active compound in the intestine. Thecomposition may also be formulated in combination with an antacid orother such ingredient.

When the dosage unit form is a capsule, it can contain, in addition tomaterial of the above type, a liquid carrier such as a fatty oil. Inaddition, dosage unit forms can contain various other materials whichmodify the physical form of the dosage unit, for example, coatings ofsugar and other enteric agents. The compounds can also be administeredas a component of an elixir, suspension, syrup, wafer, sprinkle, chewinggum or the like. A syrup may contain, in addition to the activecompounds, sucrose as a sweetening agent and certain preservatives, dyesand colorings and flavors.

The active materials can also be mixed with other active materials whichdo not impair the desired action, or with materials that supplement thedesired action, such as antacids, H2 blockers, and diuretics. The activeingredient is a compound or pharmaceutically acceptable derivativethereof as described herein. Higher concentrations, up to about 98% byweight of the active ingredient may be included.

In all embodiments, tablets and capsules formulations may be coated asknown by those of skill in the art in order to modify or sustaindissolution of the active ingredient. Thus, for example, they may becoated with a conventional enterically digestible coating, such asphenylsalicylate, waxes and cellulose acetate phthalate.

b. Liquid Compositions for Oral Administration

Liquid oral dosage forms include aqueous solutions, emulsions,suspensions, solutions and/or suspensions reconstituted fromnon-effervescent granules and effervescent preparations reconstitutedfrom effervescent granules. Aqueous solutions include, for example,elixirs and syrups. Emulsions are either oil-in-water or water-in-oil.

Elixirs are clear, sweetened, hydroalcoholic preparations.Pharmaceutically acceptable carriers used in elixirs include solvents.Syrups are concentrated aqueous solutions of a sugar, for example,sucrose, and may contain a preservative. An emulsion is a two-phasesystem in which one liquid is dispersed in the form of small globulesthroughout another liquid. Pharmaceutically acceptable carriers used inemulsions are non-aqueous liquids, emulsifying agents and preservatives.Suspensions use pharmaceutically acceptable suspending agents andpreservatives. Pharmaceutically acceptable substances used innon-effervescent granules, to be reconstituted into a liquid oral dosageform, include diluents, sweeteners and wetting agents. Pharmaceuticallyacceptable substances used in effervescent granules, to be reconstitutedinto a liquid oral dosage form, include organic acids and a source ofcarbon dioxide. Coloring and flavoring agents are used in all of theabove dosage forms.

Solvents include glycerin, sorbitol, ethyl alcohol and syrup. Examplesof preservatives include glycerin, methyl and propylparaben, benzoicacid, sodium benzoate and alcohol. Examples of non-aqueous liquidsutilized in emulsions include mineral oil and cottonseed oil. Examplesof emulsifying agents include gelatin, acacia, tragacanth, bentonite,and surfactants such as polyoxyethylene sorbitan monooleate. Suspendingagents include sodium carboxymethylcellulose, pectin, tragacanth, Veegumand acacia. Sweetening agents include sucrose, syrups, glycerin andartificial sweetening agents such as saccharin. Wetting agents includepropylene glycol monostearate, sorbitan monooleate, diethylene glycolmonolaurate and polyoxyethylene lauryl ether. Organic acids includecitric and tartaric acid. Sources of carbon dioxide include sodiumbicarbonate and sodium carbonate. Coloring agents include any of theapproved certified water soluble FD and C dyes, and mixtures thereof.Flavoring agents include natural flavors extracted from plants suchfruits, and synthetic blends of compounds which produce a pleasant tastesensation.

For a solid dosage form, the solution or suspension, in for examplepropylene carbonate, vegetable oils or triglycerides, is in oneembodiment encapsulated in a gelatin capsule. Such solutions, and thepreparation and encapsulation thereof, are disclosed in U.S. Pat. Nos.4,328,245; 4,409,239; and 4,410,545. For a liquid dosage form, thesolution, e.g., for example, in a polyethylene glycol, may be dilutedwith a sufficient quantity of a pharmaceutically acceptable liquidcarrier, e.g., water, to be easily measured for administration.

Alternatively, liquid or semi-solid oral formulations may be prepared bydissolving or dispersing the active compound or salt in vegetable oils,glycols, triglycerides, propylene glycol esters (e.g., propylenecarbonate) and other such carriers, and encapsulating these solutions orsuspensions in hard or soft gelatin capsule shells. Other usefulformulations include those set forth in U.S. Pat. Nos. RE28,819 and4,358,603. Briefly, such formulations include, but are not limited to,those containing a compound provided herein, a dialkylated mono- orpoly-alkylene glycol, including, but not limited to,1,2-dimethoxymethane, diglyme, triglyme, tetraglyme, polyethyleneglycol-350-dimethyl ether, polyethylene glycol-550-dimethyl ether,polyethylene glycol-750-dimethyl ether wherein 350, 550 and 750 refer tothe approximate average molecular weight of the polyethylene glycol, andone or more antioxidants, such as butylated hydroxytoluene (BHT),butylated hydroxyanisole (BHA), propyl gallate, vitamin E, hydroquinone,hydroxycoumarins, ethanolamine, lecithin, cephalin, ascorbic acid, malicacid, sorbitol, phosphoric acid, thiodipropionic acid and its esters,and dithiocarbamates.

Other formulations include, but are not limited to, aqueous alcoholicsolutions including a pharmaceutically acceptable acetal. Alcohols usedin these formulations are any pharmaceutically acceptable water-misciblesolvents having one or more hydroxyl groups, including, but not limitedto, propylene glycol and ethanol. Acetals include, but are not limitedto, di(lower alkyl) acetals of lower alkyl aldehydes such asacetaldehyde diethyl acetal.

2. Injectables, Solutions and Emulsions

Parenteral administration, in one embodiment characterized by injection,either subcutaneously, intramuscularly or intravenously is alsocontemplated herein. Injectables can be prepared in conventional forms,either as liquid solutions or suspensions, solid forms suitable forsolution or suspension in liquid prior to injection, or as emulsions.The injectables, solutions and emulsions also contain one or moreexcipients. Suitable excipients are, for example, water, saline,dextrose, glycerol or ethanol. In addition, if desired, thepharmaceutical compositions to be administered may also contain minoramounts of non-toxic auxiliary substances such as wetting or emulsifyingagents, pH buffering agents, stabilizers, solubility enhancers, andother such agents, such as for example, sodium acetate, sorbitanmonolaurate, triethanolamine oleate and cyclodextrins.

Implantation of a slow-release or sustained-release system, such that aconstant level of dosage is maintained (see, e.g., U.S. Pat. No.3,710,795) is also contemplated herein. Briefly, a compound providedherein is dispersed in a solid inner matrix, e.g.,polymethylmethacrylate, polybutylmethacrylate, plasticized orunplasticized polyvinylchloride, plasticized nylon, plasticizedpolyethyleneterephthalate, natural rubber, polyisoprene,polyisobutylene, polybutadiene, polyethylene, ethylene-vinylacetatecopolymers, silicone rubbers, polydimethylsiloxanes, silicone carbonatecopolymers, hydrophilic polymers such as hydrogels of esters of acrylicand methacrylic acid, collagen, cross-linked polyvinylalcohol andcross-linked partially hydrolyzed polyvinyl acetate, that is surroundedby an outer polymeric membrane, e.g., polyethylene, polypropylene,ethylene/propylene copolymers, ethylene/ethyl acrylate copolymers,ethylene/vinylacetate copolymers, silicone rubbers, polydimethylsiloxanes, neoprene rubber, chlorinated polyethylene, polyvinylchloride,vinylchloride copolymers with vinyl acetate, vinylidene chloride,ethylene and propylene, ionomer polyethylene terephthalate, butyl rubberepichlorohydrin rubbers, ethylene/vinyl alcohol copolymer,ethylene/vinyl acetate/vinyl alcohol terpolymer, andethylene/vinyloxyethanol copolymer, that is insoluble in body fluids.The compound diffuses through the outer polymeric membrane in a releaserate controlling step. The percentage of active compound contained insuch parenteral compositions is highly dependent on the specific naturethereof, as well as the activity of the compound and the needs of thesubject.

Parenteral administration of the compositions includes intravenous,subcutaneous and intramuscular administrations. Preparations forparenteral administration include sterile solutions ready for injection,sterile dry soluble products, such as lyophilized powders, ready to becombined with a solvent just prior to use, including hypodermic tablets,sterile suspensions ready for injection, sterile dry insoluble productsready to be combined with a vehicle just prior to use and sterileemulsions. The solutions may be either aqueous or nonaqueous.

If administered intravenously, suitable carriers include physiologicalsaline or phosphate buffered saline (PBS), and solutions containingthickening and solubilizing agents, such as glucose, polyethyleneglycol, and polypropylene glycol and mixtures thereof.

Pharmaceutically acceptable carriers used in parenteral preparationsinclude aqueous vehicles, nonaqueous vehicles, antimicrobial agents,isotonic agents, buffers, antioxidants, local anesthetics, suspendingand dispersing agents, emulsifying agents, sequestering or chelatingagents and other pharmaceutically acceptable substances.

Examples of aqueous vehicles include Sodium Chloride Injection, RingersInjection, Isotonic Dextrose Injection, Sterile Water Injection,Dextrose and Lactated Ringers Injection. Nonaqueous parenteral vehiclesinclude fixed oils of vegetable origin, cottonseed oil, corn oil, sesameoil and peanut oil. Antimicrobial agents in bacteriostatic orfungistatic concentrations must be added to parenteral preparationspackaged in multiple-dose containers which include phenols or cresols,mercurials, benzyl alcohol, chlorobutanol, methyl and propylp-hydroxybenzoic acid esters, thimerosal, benzalkonium chloride andbenzethonium chloride. Isotonic agents include sodium chloride anddextrose. Buffers include phosphate and citrate. Antioxidants includesodium bisulfate. Local anesthetics include procaine hydrochloride.Suspending and dispersing agents include sodium carboxymethylcelluose,hydroxypropyl methylcellulose and polyvinylpyrrolidone. Emulsifyingagents include Polysorbate 80 (TWEEN® 80). A sequestering or chelatingagent of metal ions include EDTA. Pharmaceutical carriers also includeethyl alcohol, polyethylene glycol and propylene glycol for watermiscible vehicles; and sodium hydroxide, hydrochloric acid, citric acidor lactic acid for pH adjustment.

The concentration of the pharmaceutically active compound is adjusted sothat an injection provides an effective amount to produce the desiredpharmacological effect. The exact dose depends on the age, weight andcondition of the patient or animal as is known in the art.

The unit-dose parenteral preparations are packaged in an ampoule, a vialor a syringe with a needle. All preparations for parenteraladministration must be sterile, as is known and practiced in the art.

Illustratively, intravenous or intraarterial infusion of a sterileaqueous solution containing an active compound is an effective mode ofadministration. Another embodiment is a sterile aqueous or oily solutionor suspension containing an active material injected as necessary toproduce the desired pharmacological effect.

Injectables are designed for local and systemic administration. In oneembodiment, a therapeutically effective dosage is formulated to containa concentration of at least about 0.1% w/w up to about 90% w/w or more,in certain embodiments more than 1% w/w of the active compound to thetreated tissue(s).

The compound may be suspended in micronized or other suitable form ormay be derivatized to produce a more soluble active product or toproduce a prodrug. The form of the resulting mixture depends upon anumber of factors, including the intended mode of administration and thesolubility of the compound in the selected carrier or vehicle. Theeffective concentration is sufficient for ameliorating the symptoms ofthe condition and may be empirically determined.

3. Lyophilized Powders

Of interest herein are also lyophilized powders, which can bereconstituted for administration as solutions, emulsions and othermixtures. They may also be reconstituted and formulated as solids orgels.

The sterile, lyophilized powder is prepared by dissolving a compoundprovided herein, or a pharmaceutically acceptable derivative thereof, ina suitable solvent. The solvent may contain an excipient which improvesthe stability or other pharmacological component of the powder orreconstituted solution, prepared from the powder. Excipients that may beused include, but are not limited to, dextrose, sorbital, fructose, cornsyrup, xylitol, glycerin, glucose, sucrose or other suitable agent. Thesolvent may also contain a buffer, such as citrate, sodium or potassiumphosphate or other such buffer known to those of skill in the art at, inone embodiment, about neutral pH. Subsequent sterile filtration of thesolution followed by lyophilization under standard conditions known tothose of skill in the art provides the desired formulation. In oneembodiment, the resulting solution will be apportioned into vials forlyophilization. Each vial will contain a single dosage or multipledosages of the compound. The lyophilized powder can be stored underappropriate conditions, such as at about 4° C. to room temperature.

Reconstitution of this lyophilized powder with water for injectionprovides a formulation for use in parenteral administration. Forreconstitution, the lyophilized powder is added to sterile water orother suitable carrier. The precise amount depends upon the selectedcompound. Such amount can be empirically determined.

4. Topical Administration

Topical mixtures are prepared as described for the local and systemicadministration. The resulting mixture may be a solution, suspension,emulsions or the like and are formulated as creams, gels, ointments,emulsions, solutions, elixirs, lotions, suspensions, tinctures, pastes,foams, aerosols, irrigations, sprays, suppositories, bandages, dermalpatches or any other formulations suitable for topical administration.

The compounds or pharmaceutically acceptable derivatives thereof may beformulated as aerosols for topical application, such as by inhalation(see, e.g., U.S. Pat. Nos. 4,044,126, 4,414,209, and 4,364,923, whichdescribe aerosols for delivery of a steroid useful for treatment ofinflammatory diseases, particularly asthma). These formulations foradministration to the respiratory tract can be in the form of an aerosolor solution for a nebulizer, or as a micro fine powder for insufflation,alone or in combination with an inert carrier such as lactose. In such acase, the particles of the formulation will, in one embodiment, havediameters of less than 50 microns, in one embodiment less than 10microns.

The compounds may be formulated for local or topical application, suchas for topical application to the skin and mucous membranes, such as inthe eye, in the form of gels, creams, and lotions and for application tothe eye or for intracisternal or intraspinal application. Topicaladministration is contemplated for transdermal delivery and also foradministration to the eyes or mucosa, or for inhalation therapies. Nasalsolutions of the active compound alone or in combination with otherpharmaceutically acceptable excipients can also be administered.

For nasal administration, the preparation may contain an esterifiedphosphonate compound dissolved or suspended in a liquid carrier, inparticular, an aqueous carrier, for aerosol application. The carrier maycontain solubilizing agents such as propylene glycol, surfactants,absorption enhancers such as lecithin or cyclodextrin, or preservatives.

These solutions, particularly those intended for ophthalmic use, may beformulated as 0.01%-10% isotonic solutions, pH about 5-7, withappropriate salts.

5. Compositions for Other Routes of Administration

Other routes of administration, such as transdermal patches, includingiontophoretic and electrophoretic devices, and rectal administration,are also contemplated herein.

Transdermal patches, including iotophoretic and electrophoretic devices,are well known to those of skill in the art. For example, such patchesare disclosed in U.S. Pat. Nos. 6,267,983, 6,261,595, 6,256,533,6,167,301, 6,024,975, 6,010,715, 5,985,317, 5,983,134, 5,948,433, and5,860,957.

For example, pharmaceutical dosage forms for rectal administration arerectal suppositories, capsules and tablets for systemic effect. Rectalsuppositories are used herein mean solid bodies for insertion into therectum which melt or soften at body temperature releasing one or morepharmacologically or therapeutically active ingredients.Pharmaceutically acceptable substances utilized in rectal suppositoriesare bases or vehicles and agents to raise the melting point. Examples ofbases include cocoa butter (theobroma oil), glycerin-gelatin, carbowax(polyoxyethylene glycol) and appropriate mixtures of mono-, di- andtriglycerides of fatty acids. Combinations of the various bases may beused. Agents to raise the melting point of suppositories includespermaceti and wax. Rectal suppositories may be prepared either by thecompressed method or by molding. The weight of a rectal suppository, inone embodiment, is about 2 to 3 gm.

Tablets and capsules for rectal administration are manufactured usingthe same pharmaceutically acceptable substance and by the same methodsas for formulations for oral administration.

6. Targeted Formulations

The compounds provided herein, or pharmaceutically acceptablederivatives thereof, may also be formulated to be targeted to aparticular tissue, receptor, or other area of the body of the subject tobe treated. Many such targeting methods are well known to those of skillin the art. All such targeting methods are contemplated herein for usein the instant compositions. For non-limiting examples of targetingmethods, see, e.g., U.S. Pat. Nos. 6,316,652, 6,274,552, 6,271,359,6,253,872, 6,139,865, 6,131,570, 6,120,751, 6,071,495, 6,060,082,6,048,736, 6,039,975, 6,004,534, 5,985,307, 5,972,366, 5,900,252,5,840,674, 5,759,542 and 5,709,874.

In one embodiment, liposomal suspensions, including tissue-targetedliposomes, such as tumor-targeted liposomes, may also be suitable aspharmaceutically acceptable carriers. These may be prepared according tomethods known to those skilled in the art. For example, liposomeformulations may be prepared as described in U.S. Pat. No. 4,522,811.Briefly, liposomes such as multilamellar vesicles (MLV's) may be formedby drying down egg phosphatidyl choline and brain phosphatidyl serine(7:3 molar ratio) on the inside of a flask. A solution of a compoundprovided herein in phosphate buffered saline lacking divalent cations(PBS) is added and the flask shaken until the lipid film is dispersed.The resulting vesicles are washed to remove unencapsulated compound,pelleted by centrifugation, and then resuspended in PBS.

7. Articles of Manufacture

The compounds or pharmaceutically acceptable derivatives may be packagedas articles of manufacture containing packaging material, a compound orpharmaceutically acceptable derivative thereof provided herein, which iseffective for treatment, prevention or amelioration of one or moresymptoms of diseases or disorders associated with viral infections,inappropriate cell proliferation or bone metabolism, within thepackaging material, and a label that indicates that the compound orcomposition, or pharmaceutically acceptable derivative thereof, is usedfor the treatment, prevention or amelioration of one or more symptoms ofdiseases or disorders associated with viral infections, inappropriatecell proliferation or bone metabolism.

The articles of manufacture provided herein contain packaging materials.Packaging materials for use in packaging pharmaceutical products arewell known to those of skill in the art. See, e.g., U.S. Pat. Nos.5,323,907, 5,052,558 and 5,033,252. Examples of pharmaceutical packagingmaterials include, but are not limited to, blister packs, bottles,tubes, inhalers, pumps, bags, vials, containers, syringes, bottles, andany packaging material suitable for a selected formulation and intendedmode of administration and treatment. A wide array of formulations ofthe compounds and compositions provided herein are contemplated as are avariety of treatments for any disease or disorder associated with viralinfections, inappropriate cell proliferation or bone metabolism.

E. Evaluation of the Activity of the Compounds

The activity of the compounds as antivirals can be measured in standardassays known in the art. Exemplary assays are described herein.

Determination of Antiviral Activity and Drug Cytotoxicity

(VV and CV Activity)

The tables below show CC₅₀ and EC₅₀ data for exemplary compounds. InTable 1, antiviral activity and cytotoxicity of compounds providedherein against vaccinia virus (VV) and cowpox virus (CV) in humanforeskin fibroblast (HFF) cells infected with VV or CV in plaquereduction assay and neutral red uptake assay known in the art wasmeasured. Eicosyl-(S)-HPMPA and 15-methyl-hexadecyloxyethyl-(S)-HPMPAshow EC₅₀ less than 0.5 μM in this assay.

TABLE 1 Efficacy and Cytotoxicity of alkyl esters of Cidofovir (CDV) andcyclic Cidofovir (cCDV) in Vaccinia Copenhagen and Cowpox BrightonVaccinia Copenhagen Cowpox Brighton Compound Abbreviation CC₅₀ (μM)^(a)EC₅₀ (μM)^(a) SI^(b) EC₅₀ (μM)^(a) SI^(b) Cidofovir Series CidofovirCDV >317 ± 0  31 ± 5.4 >10  42 ± 5.4 >7.5 R group Octyl- (8)O-CDV >100 >20 ± 0 — >20 ± 0 — Dodecyl- (12) DD-CDV >100 >20 ± 0 — >20 ±0 — Hexadecyl- (16) HD-CDV >157 ± 58  3.1 ± 0.1 >51  6.0 ± 2.0 >26Eicosyl- (20) EC-CDV  45 ± 8.5  1.6 ± 1.3 28  1.5 ± 0.9 30 Tetracosyl-(24) TC-CDV >100 >17 ± 4.2 — >20 ± 0 — ^(a)Values are the means of twoor more assays (± standard deviation) ^(b)SI = CC₅₀/EC₅₀ ^(c)Values inparentheses are the numbers of atoms beyond the phosphate oxygen.HCMV and MCMV Activityi) Cell Cultures and Viruses

Human foreskin fibroblast (HFF) cells and mouse embryo fibroblast (MEF)cells are prepared as primary cultures and used in the HCMV and MCMVassays. Cells are propagated in minimal essential medium (MEM)containing 10% fetal bovine serum (FBS), 2 mM L-glutamine, 100 U ofpenicillin per ml and 25 μg of gentamicin per ml in T-175 cm2 tissueculture flasks (BD Falcon, Bedford, Mass.) until used in antiviralassays. HCMV strain AD-169 and MCMV strain Smith are propagated by usingstandard virological techniques.

(ii) Neutral Red Uptake Assay for Cytotoxicity.

HFF cells are seeded into 96-well tissue culture plates at 2.5×104cells/well. After a 24-h incubation, medium is replaced with MEMcontaining 2% FBS, and an alkyl ester provided herein is added to thefirst row and then diluted serially fivefold from 100 to 0.03 μM. Theplates are incubated for 7 days, and cells are stained with neutral redand incubated for 1 h. Plates are shaken on a plate shaker for 15 min,and neutral red is solubilized with 1% glacial acetic acid-50% ethanol.The optical density is read at 540 nm. The concentration of drug thatreduced cell viability by 50% (CC₅₀) is calculated by using computersoftware. MEF cells are stained with neutral red and evaluated visuallywith a stereomicroscope at ×10 magnification. No toxicity was observedin the MCMV assays at the concentrations tested. The CC₅₀ data forexemplary compounds is shown in Tables 1 and 2.

(iii) Plaque Reduction Assay for Antiviral Activity.

HFF or MEF cells are placed into 6- or 12-well plates and incubated at37° C. for 2 days (HFF) or 1 day (MEF). Ganciclovir and CDV are used aspositive controls. Virus is diluted in MEM containing 10% FBS to provide20 to 30 plaques per well. The medium is aspirated, and 0.2 ml of virusis added to each well in triplicate with 0.2 ml of medium added to drugtoxicity wells. The plates are incubated for 1 h with shaking every 15min. An alkyl ester provided herein is serially diluted 1:5 in MEM with2% FBS starting at 100 μM and added to appropriate wells. Following a7-day incubation for MCMV, or 8 days for HCMV, cells are strained for 6h with 2 ml of 5.0% neutral red in phosphate-buffered saline. The stainis aspirated, and plaques were counted by using a stereomicroscope at×10 magnification. By comparing drug-treated with untreated wells, 50%effective concentrations (EC₅₀s) are calculated in a standard manner.The EC₅₀ data for exemplary compounds is shown in Table 2.

TABLE 2 Efficacy and Cytotoxicity of alkyl esters of Cidofovir (CDV) andcyclic Cidofovir (cCDV) for HCMV and MCMV HCMV MCMV EC₅₀ EC₅₀ CompoundAbbreviation CC₅₀ (μM)^(a) (μM)^(a) SI (μM)^(a) SI Cidofovir SeriesCidofovir CDV >317 ± 0       1.2 ± 0.43 264 0.04 ± 0.03 >7925 R groupOctyl- (8:0) O-CDV >100     5.25 ± 5.7 >19 >10 <10 Dodecyl-(12:0)DD-CDV >100     0.31 ± 0.19 >322 >10 <10 Hexadecyl- (16:0) HD-CDV >157 ±58   0.005 ± 0 >31400 0.02 ± 0   >7850 Eicosyl- (20:0) EC-CDV 38.5 0.0007 5500 0.005 ± 0    7700 Docosyl- (22:0) DC-CDV 79.6 0.004 >26500 0.85 ± 0.007 995 Tetracosyl- (24:0) TC-CDV >100     0.006 ± 0.03 16707.45 ± 0.64 >13 Cyclic Cidofovir Series Cyclic Cidofovir cCDV >331 ± 0     3.1 ± 1.8 >107 0.23 ± 0.07 >1439 R group Octyl- (8:0) O-cCDV Nd NdNd Nd Nd Dodecyl-(12:0) DD-cCDV Nd Nd Nd Nd Nd Hexadecyl- HD-cCDV  83 ±20     3.10 ± 2.35 26.8 0.13 ± 0.01 6638 (16:0) Eicosyl- (20:0) EC-cCDV86.1 0.011 7827 0.23 ± 0.01 374 Docosyl- (22:0) DC-cCDV >100 0.032 >31252.8 ± 1.7 >36 Tetracosyl- TC-cCDV >100 0.32  >313 2.3 ± 1.3 >43 (24:0)Nd = not determined ^(a)Values are the means of two or more assays(±standard deviation) ^(b)SI = CC₅₀/EC₅₀ Values in parentheses are thenumbers of atoms beyond the phosphate oxygen; the number after the colonis the number of double bonds in the alkyl chain. Data are recorded asthe means ± standard deviations of at least two determinations. Valueswithout a standard deviation represent a single determination. SI,selective index (CC₅₀/EC₅₀).

F. Methods of Use of the Compounds and Compositions

Methods of treating, preventing, or ameliorating one or more symptoms ofdiseases associated with viral infections, inappropriate cellproliferation and bone metabolism using the compounds and compositionsprovided herein are provided. In practicing the methods, effectiveamounts of the compounds or compositions containing therapeuticallyeffective concentrations of the compounds are administered. In certainembodiments, the methods provided herein are for the preventing, orameliorating one or more symptoms of diseases associated with viralinfections, including, but not limited to influenza; hepatitis B and Cvirus; cytomegalovirus (CMV); herpes infections, such as those caused byVaricella zoster virus, Herpes simplex virus types 1 & 2, Epstein-Barrvirus, Herpes type 6 (HHV-6) and type 8 (HHV-8); Varicella zoster virusinfections such as shingles or chicken pox; Epstein Barr virusinfections, including, but not limited to infectiousmononucleosis/glandular; retroviral infections including, but notlimited to SIV, HIV-1 and HIV-2; ebola virus; adenovirus and papillomavirus.

In further embodiments, the methods provided herein are for thepreventing, treating, or ameliorating one or more symptoms of diseasesassociated with viral infections caused by orthopox viruses, such asvariola major and minor, vaccinia, smallpox, cowpox, camelpox, andmonkeypox.

In one embodiment, a therapeutically effective dosage to treat such anorthopox infection should produce a serum concentration of activeingredient of from about 0.1 ng/ml to about 50-100 μg/ml. Thepharmaceutical compositions, in another embodiment, should provide adosage of from about 0.001 mg to about 2000 mg of compound per kilogramof body weight per day. Pharmaceutical dosage unit forms are prepared,e.g., to provide from about 0.01 mg, 0.1 mg or 1 mg to about 500 mg,1000 mg or 2000 mg, and in one embodiment from about 10 mg to about 500mg of the active ingredient or a combination of essential ingredientsper dosage unit form.

In certain embodiments, the methods provided herein are for thepreventing, or ameliorating one or more symptoms of diseases associatedwith cell proliferation, including, but not limited to cancers. Examplesof cancers include, but are not limited to, lung cancer, head and necksquamous cancers, colorectal cancer, prostate cancer, breast cancer,acute lymphocytic leukemia, adult acute myeloid leukemia, adult nonHodgkin's lymphoma, brain tumors, cervical cancers, childhood cancers,childhood sarcoma, chronic lymphocytic leukemia, chronic myeloidleukemia, esophageal cancer, hairy cell leukemia, kidney cancer, livercancer, multiple myeloma, neuroblastoma, oral cancer, pancreatic cancer,primary central nervous system lymphoma, and skin cancer.

Methods of treating, preventing, or ameliorating one or more symptomsassociated with bone metabolism by administering a compound orcomposition provided herein are provided. Such diseases include, but arenot limited to osteoporosis, such as senile, post-menopausal orsteroid-induced osteoporosis, metastatic bone cancers, Paget's disease,osteogenesis imperfecta, fibrous dysplasia, rheumatoid arthritis,hyperparathyroidism, algodystrophy, sterno-costoclavicular hyperostosis,Gaucher's disease, Engleman's disease, certain non-skeletal disordersand periodontal disease. An example of an effective amount is an amountthat will prevent, attenuate, or reverse abnormal or excessive boneresorption or the bone resorption that occurs in the aged, particularlypost-menopausal females or prevent or oppose bone metastasis andvisceral metastasis in breast cancer.

G. Combination Therapy

The compounds and compositions provided herein may also be used incombination with other active ingredients. In certain embodiments, thecompounds may be administered in combination, or sequentially, withanother therapeutic agent. Such other therapeutic agents include thoseknown for treatment, prevention, or amelioration of one or more symptomsassociated with viral infections, inappropriate cell proliferation andbone metabolism. Such therapeutic agents include, but are not limitedto, antiviral agents, anti-neoplastic agents and agents for thetreatment and/or prevention of symptoms associated bone metabolism. Incertain embodiments, the compounds provided herein may be administeredin combination with one or more antiviral agents or anti-cancer agents.Anti-cancer agents for use in combination with the instant compoundsinclude, but are not limited to, alkylating agents, toxins,antiproliferative agents and tubulin binding agents. Classes ofcytotoxic agents for use herein include, for example, the anthracyclinefamily of drugs, the vinca drugs, the mitomycins, the bleomycins, thecytotoxic nucleosides, the pteridine family of drugs, diynenes, themaytansinoids, the epothilones, the taxanes and the podophyllotoxins.

It should be understood that any suitable combination of the compoundsprovided herein with one or more of the above-mentioned compounds andoptionally one or more further pharmacologically active substances areconsidered to be within the scope of the present disclosure. In anotherembodiment, the compound provided herein is administered prior to orsubsequent to the one or more additional active ingredients.

The following examples are provided for illustrative purposes only andare not intended to limit the scope of the invention.

EXAMPLES

1H and 31P nuclear magnetic resonance (NMR) spectra were recorded on aVarian HG-400 spectrometer with tetramethylsilane (internal) and 85%D3PO4 in D20 (external) as references for 1H and 31P (0.00 ppm),respectively. Electrospray ionization mass spectroscopy (ESI) wasperformed by HT Laboratories (San Diego, Calif.). Chromatographicpurification was done by the flash method with Merck silica gel 60 (240to 400 mesh). All final products were homogeneous by thin-layerchromatography performed on Analtech 250-μm Silica Gel GF Uniplatesvisualized under UV light, with phospray (Supelco, Bellafonte, Pa.), andby charring (400° C.). Cyclic cidofovir dihydrate was prepared by themethod of Louie and Chapman (Nucleosides & Nucleic Acids, 20:1099-1102,2001). Bromoalkanes and bromoalkoxyalkanes were either commerciallyavailable or synthesized from the corresponding alcohol. All otherchemicals were of reagent quality and used as obtained from thesuppliers. All reactions were carried out in an inert atmosphere (drynitrogen).

Example 1 Octyl Cyclic Cidofovir

1-bromooctane (1.2 eq) was added to a suspension of cyclic cidofovir(cCDV) dihydrate, N,N-dicyclohexyl-4-morpholino-carboxamidine (1.1 eq)and N,N-dimethylformamide (10 ml/mmol). The mixture was heated to 60° C.and stirred magnetically overnight. The solvent was then evaporatedunder reduced pressure, the residue was adsorbed onto silica gel andpurified by flash column chromatography (elution gradient, CH₂Cl₂ to 15%EtOH). The Octyl cyclic cidofovir was isolated as equimolar mixtures ofthe axial and equatorial diastereomers.

Yield 46%; ¹H NMR (CDCl₃+CD₃OD) δ 7.00 (dd, J=7.4 Hz, J=10 Hz, 1H), 5.44(d, J=7.4 Hz, 1H), 3.99 to 4.29 (m, 2H), 3.64 to 3.86 (m, 6H), 3.50 to3.58 (m, 2H), 3.12 to 3.33 (m, 1H), 1.28 to 1.39 (m, 2H), 0.75 to 1.15(m, 10H), 0.52 (t, J=6.8 Hz); ³¹P NMR δ 12.31, 13.58.

MS (ESI): m/z 374 (M+H)⁺, 372 (M−H)⁻.

Example 2 Dodecyl Cyclic Cidofovir

The title compound was prepared using the same procedure as describedfor Example 1 using 1-bromododecane and cyclic cidofovir.

Yield 45%; ¹H NMR (CDCl₃+CD₃OD) δ 7.02 (dd, J=7.1 Hz, J=10 Hz, 1H), 5.47(d, J=7.4 Hz, 1H), 4.03 to 4.07 (m, 2H), 3.69 to 3.89 (m, 6H), 3.53 to3.60 (m, 2H), 3.15 to 3.35 (m, 1H), 1.32 to 1.42 (m, 2H), 0.86 to 1.15(m, 10H), 0.54 (t, J=6.0 Hz); ³¹P NMR (CDCl₃+CD₃OD) δ=1.2.22, 13.51. MS(ESI): m/z 430 (M+H)⁺, 452 (M+Na)⁺; 428 (M−H)⁻.

Example 3 Hexadecyl Cyclic Cidofovir

The title compound was prepared using the same procedure as describedfor Example 1 using 1-bromohexadecane and cyclic cidofovir.

Yield 55%; ¹H NMR (di-methyl sulfoxide [DMSO]-d₆) δ 7.48 (dd, J=31.2,7.2 Hz, 1H), 7.108 (br d, J=39.9 Hz, 2H), 5.64 (t, J=6.8 Hz, 1H), 3.5 to4.9 (m, 7H), 1.60 (br s, 2H), 1.24 (br s, 28H), 0.86 (t, J=6.5 Hz, 3H);³¹P NMR (CDCl₃+CD₃OD) δ=12.18, 13.58. MS (ESI): m/z 486 (MH)⁺, 484(M−H)⁻.

Example 4 Eicosyl Cyclic Cidofovir (EC-cCDV)

The title compound was prepared using the same procedure as describedfor Example 1 using 1-bromoeicosane and cyclic cidofovir.

Yield 8%; ¹H NMR (DMSO-d₆) δ 7.43 (d, J=7.2 Hz, 1H), 7.08 (br d, J=30.9Hz 2H), 5.61 (d, J=6.9 Hz, 1H), 3.5 to 4.9 (m, 7H), 1.60 (br s, 2H),1.24 (br s, 36H), 0.86 (t, J=6.5 Hz, 3H); ³¹P NMR δ=14.02, 12.81; MS(ESI): m/z 564 (M+Na)⁺, 542 (M+H)⁺.

Example 5 Tetracosyl Cyclic Cidofovir

The title compound was prepared using the same procedure as describedfor Example 1 using 1-bromotetracosane and cyclic cidofovir.

Yield 17%; ¹H NMR (DMSO-d₆) δ 7.43 (d, J=6.9 Hz, 1H), 7.07 (br d, J=25.5Hz 2H), 5.63 (d, J=6.9 Hz, 1H), 3.1 to 4.0 (m, 7H), 1.60 (br s, 2H),1.22 (br s, 44H), 0.84 (t, J=6.5 Hz, 3H); ³¹P NMR (CDCl₃+CD₃OD) δ 12.66,13.86. MS (ESI): m/z 620 (M+Na)⁺, 598 (M+H)⁺.

Example 6 Docosyl Cyclic Cidofovir

The title compound was prepared by Mitsunobu reaction. Anhydrous cCDV (1eq), the docosanol (2 eq), and tri-phenylphosphine (2 eq) were dissolvedor suspended in anhydrous N,N-dimethylformamide (6.5 ml per mmol ofcCDV) and stirred vigorously under a nitrogen atmosphere. Diisopropylazadicarboxylate (2 eq) was then added in three portions over 15 minbefore the mixture was allowed to stir overnight. The solvent was thenevaporated under vacuum, and the residue was adsorbed onto silica geland purified by column chromatography. Gradient elution from 100% CH₂Cl₂to 15% EtOH was followed by recrystallization from p-dioxane. Thecoupled product was isolated as equimolar mixtures of axial andequatorial diastereomers.

Yield 26%; ¹H NMR (DMSOd₆) δ 7.43 (d, J=7.2 Hz, 1H), 7.07 (br d, J=25.5Hz 2H), 5.61 (d, J=6.9 Hz, 1H), 3.3 to 4.4 (m, 7H), 1.60 (br s, 2H),1.23 (br s, 40H), 0.86 (t, J=6.5 Hz, 3H); ³¹P NMR δ 12.80; MS (ESI): m/z564 (M+Na)⁺, 570 (M+H)⁺, 604 (MCl)⁻, 568 (M−H)⁻.

Example 7 1-O-Octadecyl-2-O-benzyl-sn-glycero-3-cCDV (ODBGcCDV)

Yield 45%; ¹H NMR (CDCl₃) δ 7.27 to 7.38 (m, 7H), 7.16 and 7.30 (pair ofdoublets, 1H), 5.72 and 5.68 (pair of doublets, 1H), 4.68 and 4.62 (pairof singlets, 2H), 3.97 to 4.40 (m, 9H), 3.44 (t, 2H), 3.41 (t, 2H), 3.25(m, 1H), 1.54 (m, 2H), 1.26 (br s, 30H), 0.88 (t, 3H); ³¹P NMR δ13.72and 12.01; MS (ESI) m/z 678 (M+H)⁺, 700 (M+Na)⁺, 676 (M−H)⁻.

Example 8 Octyl Cidofovir, Sodium Salt

The octyl cyclic CDVanalog was suspended in 2 M NaOH (25 ml/mmol). Thesuspensions was heated to 80° C. and stirred for 1 h, during which themixtures became clear. After hydrolysis, the solution was cooled to 25°C. and acidified with glacial acetic acid (pH approximately 5). Theresulting precipitate was collected by vacuum filtration and dried undervacuum. The crude product was purified either by flash columnchromatography (eluant, CH₂Cl₂-20% MeOH) and recrystallized to purityfrom ethanol (2).

Yield 46%; ¹H NMR (DMSO-d₆) δ 7.53 (d, J=7.5 Hz, 1H), 7.12 (br d, J=58.5Hz 2H), 5.64 (d, J=7.5 Hz, 1H), 3.20 to 3.80 (m, 7H), 1.42 (br s, 2H),1.24 (br s, 12H), 0.85 (t, J=6.5 Hz, 3H); ³¹P NMR δ 14.46; MS (ESI): m/z436 (M+2Na)⁺, 414 (M+Na)⁺; 390 (M−H)⁻.

The following acyclic analogs were prepared using similar procedure.

Example 9 Dodecyl Cidofovir, Sodium Salt

Yield 32%; ¹H NMR (DMSO-d₆) 67.54 (d, J=7.2 Hz, 1H), 7.19 (br d, J=57.3Hz 2H), 5.67 (d, J=7.2 Hz, 1H), 3.20 to 3.81 (m, 7H), 1.42 (br s, 2H),1.22 (br s, 20H), 0.84 (t, J=6.5 Hz, 3H); ³¹P NMR δ 14.60; MS (ESI): m/z492 (M+2Na)⁺, 470 (M+Na)⁺; 446 (M−H)⁻.

Example 10 Hexadecyl Cidofovir, Sodium Salt

Yield 43%; ¹H NMR (DMSO-d) 67.52 (d, J=7.2 Hz, 1H), 7.19 (br d, J=57.3Hz 2H), 5.65 (d, J=7.2 Hz, 1H), 3.2 to 3.9 (m, 7H), 1.45 (br s, 2H),1.24 (br s, 28H), 0.86 (t, J=6.5 Hz, 3H); ³¹P NMR (D₂O) δ=16.73; MS(ESI): m/z 504 (M+Na)⁺, 526 (M+2Na)⁺, 502 (M−H)⁻.

Example 11 Eicosyl Cidofovir, Sodium Salt (EC-CDV)

Yield 39%; ¹H NMR (DMSO-d⁸) δ=7.49 (d, J=7.2 Hz, 1H), 7.01 (br d, J=45.3Hz 2H), 5.60 (d, J=7.2 Hz, 1H), 3.2 to 3.9 (m, 7H), 1.42 (br s, 2H),1.26 (br s, 36H), 0.85 (t, J=6.5 Hz, 3H); ³¹P NMR δ=14.22; MS (ESI): m/z604 (M+2Na)⁺, 582 (M+Na)⁺; 558 (M−H)⁻.

Example 12 Docosyl Cidofovir, Sodium Salt

Yield 57%; ¹H NMR (DMSO-d₆) δ 7.50 (d, J=7.2 Hz, 1H), 7.03 (br d, J=50.4Hz 2H), 5.60 (d, J=7.2 Hz, 1H), 3.2 to 3.9 (m, 7H), 1.42 (br s, 2H),1.22 (br s, 40H), 0.85 (t, J=6.5 Hz, 3H); ³¹P NMR δ14.21; MS (ESI): m/z632 (M+2Na)⁺, 582 (M−H, sodium salt plus H)⁺; 540 (M−H)⁻, 576 (M−Cl).

Example 13 Tetracosyl Cidofovir, Sodium Salt

Yield 35%; ¹H NMR (CDCl₃+methanol d⁴) δ 7.19 (d, J=7.4 Hz, 1H), 5.41 (d,J=7.4 Hz, 1H), 3.03 to 3.62 (m, 7H), 2.90 to 2.91 (m, 2H), 1.10 to 1.22(m, 2H), 0.83 (br s, 42H), 0.45 (t, J=6.9 Hz). ³¹P NMR (CDCl₃+methanold⁴) δ=16.14. MS (ESI): m/z 638 (M+H, sodium salt plus H)⁺, 660 (M+Na,sodium salt of sodium salt)⁺; 614 (free acid)⁻.

Since modifications will be apparent to those of skill in the art, it isintended that the invention be limited only by the scope of the appendedclaims.

1. Eicosyl ester of cidofovir.
 2. A pharmaceutical compositioncomprising the compound of claim 1 and a pharmaceutically acceptablecarrier.
 3. A method for treating a viral infection in a mammal, whereinthe method comprises administering to a subject in need thereof aneffective amount of the compound of claim
 1. 4. The method of claim 3,wherein the viral infection is caused by influenza, hepatitis B virus,hepatitis C virus, cytomegalovirus, Varicella zoster virus, Herpessimplex virus types 1 and 2, Epstein-Barr virus, Herpes type 6 and type8, Varicella zoster virus, Epstein Barr virus infections, retroviralinfections, orthopox viruses, vaccinia, ebola virus, adenovirus andpapilloma virus.
 5. The method of claim 3, wherein the viral infectionis an orthopox viral infection selected from the group consisting ofsmallpox, cowpox, camelpox, and monkeypox.
 6. The method of claim 3,wherein the viral infection is a herpes viral infection.
 7. An articleof manufacture, comprising packaging material and the compound of claim1, contained within the packaging material, wherein the compound iseffective for treatment of a disease associated with a viral infectionand the packaging material includes a label that indicates that thecompound is used for treatment of a disease associated with a viralinfection.
 8. The method of claim 3, wherein the viral infection is ahepatitis viral infection.